U.S. patent number 11,274,321 [Application Number 15/963,536] was granted by the patent office on 2022-03-15 for use of oxyhydrogen microorganisms for non-photosynthetic carbon capture and conversion of inorganic and/or c1 carbon sources into useful organic compounds.
This patent grant is currently assigned to Kiverdi, Inc.. The grantee listed for this patent is Kiverdi, Inc.. Invention is credited to Lisa Dyson, John S. Reed.
United States Patent |
11,274,321 |
Reed , et al. |
March 15, 2022 |
Use of oxyhydrogen microorganisms for non-photosynthetic carbon
capture and conversion of inorganic and/or C1 carbon sources into
useful organic compounds
Abstract
Compositions and methods for a hybrid biological and chemical
process that captures and converts carbon dioxide and/or other
forms of inorganic carbon and/or CI carbon sources including but
not limited to carbon monoxide, methane, methanol, formate, or
formic acid, and/or mixtures containing CI chemicals including but
not limited to various syngas compositions, into organic chemicals
including biofuels or other valuable biomass, chemical, industrial,
or pharmaceutical products are provided. The present invention, in
certain embodiments, fixes inorganic carbon or CI carbon sources
into longer carbon chain organic chemicals by utilizing
microorganisms capable of performing the oxyhydrogen reaction and
the autotrophic fixation of CO.sub.2 in one or more steps of the
process.
Inventors: |
Reed; John S. (Hayward, CA),
Dyson; Lisa (Hayward, CA) |
Applicant: |
Name |
City |
State |
Country |
Type |
Kiverdi, Inc. |
Hayward |
CA |
US |
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Assignee: |
Kiverdi, Inc. (Pleasanton,
CA)
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Family
ID: |
1000006174060 |
Appl.
No.: |
15/963,536 |
Filed: |
April 26, 2018 |
Prior Publication Data
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Document
Identifier |
Publication Date |
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US 20180245108 A1 |
Aug 30, 2018 |
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Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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13643872 |
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PCT/US2011/034218 |
Apr 27, 2011 |
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PCT/US2010/001402 |
May 12, 2010 |
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12613550 |
Nov 6, 2009 |
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61111794 |
Nov 6, 2008 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C12N
1/20 (20130101); C12M 23/34 (20130101); C12N
1/205 (20210501); C12M 29/18 (20130101); C12M
29/08 (20130101); C12M 29/20 (20130101); C12P
7/6463 (20130101); C12M 29/02 (20130101); C25B
15/02 (20130101); C12M 43/04 (20130101); C12M
47/02 (20130101); C25B 1/04 (20130101); C12P
7/625 (20130101); C12N 1/12 (20130101); Y02E
60/36 (20130101); Y02P 20/133 (20151101) |
Current International
Class: |
C12N
1/20 (20060101); C12P 7/62 (20060101); C12P
7/625 (20220101); C12N 1/12 (20060101); C25B
15/02 (20210101); C12P 7/6463 (20220101); C25B
1/04 (20210101); C12M 1/00 (20060101) |
References Cited
[Referenced By]
U.S. Patent Documents
Other References
Volova et al., Appl Microbiol Biotechnol, 58:675-678, 2002 (Year:
2002). cited by examiner .
Schlegel et al., In: Advances in Biochemical Engineering, 1971,
vol. 1, pp. 143-168. (Year: 1971). cited by examiner .
DeCicco, B.T., Removal of Eutrophic Nutrients from Wastewater and
their Bioconversion to Bacterial Single Cell Protein for Animal
Feed Supplements Phase 1, Water Resources Research Center,
Washington Technical Institute, Washington, D.C. 20008, Nov. 1977.
cited by applicant .
DeCicco, B.T., Eutrophic Nutrients from Wastewater and their
Bioconversion to Bacterial Single Cell Protein for Animal Feed
Supplements Phase II, Water Resources Research Center, University
of the District of Columbia, Van Ness Campus, Washington, D.C.
20008, Apr. 1979. cited by applicant .
U.S. Appl. No. 16/013,833, Office Action, dated Apr. 18, 2019.
cited by applicant.
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Primary Examiner: Fox; Allison M
Assistant Examiner: Xu; Qing
Attorney, Agent or Firm: FisherBroyles, LLP Jacobson; Jill
A.
Parent Case Text
RELATED APPLICATIONS
This application is a continuation of U.S. application Ser. No.
13/643,872, filed Mar. 1, 2013, which is a national stage of
International Patent Application No. PCT/US2011/034218, filed Apr.
27, 2011, which claims priority under 35 U.S.C. .sctn. 119(e) to
U.S. Provisional Patent Application No. 61/328,184, filed Apr. 27,
2010 and entitled "USE OF OXYHYDROGEN MICROORGANISMS FOR
NON-PHOTOSYNTHETIC CARBON CAPTURE AND CONVERSION OF INORGANIC
CARBON SOURCES INTO USEFUL ORGANIC COMPOUNDS." International Patent
Application No. PCT/US2011/034218 is also a continuation-in-part of
International Patent Application No. PCT/US2010/001402, filed May
12, 2010, and entitled "BIOLOGICAL AND CHEMICAL PROCESS UTILIZING
CHEMOAUTOTROPHIC MICROORGANISMS FOR THE CHEMOSYNTHETIC FIXATION OF
CARBON DIOXIDE AND/OR OTHER INORGANIC CARBON SOURCES INTO ORGANIC
COMPOUNDS, AND THE GENERATION OF ADDITIONAL USEFUL PRODUCTS," which
is a continuation-in-part of U.S. patent application Ser. No.
12/613,550, filed Nov. 6, 2009, and entitled "BIOLOGICAL AND
CHEMICAL PROCESS UTILIZING CHEMOAUTOTROPHIC MICROORGANISMS FOR THE
CHEMOSYNTHETIC FIXATION OF CARBON DIOXIDE AND/OR OTHER INORGANIC
CARBON SOURCES INTO ORGANIC COMPOUNDS, AND THE GENERATION OF
ADDITIONAL USEFUL PRODUCTS," which claims the benefit of U.S.
Provisional Patent Application No. 61/111,794, filed Nov. 6, 2008,
and entitled, "BIOLOGICAL AND CHEMICAL PROCESS UTILIZING
CHEMOAUTOTROPHIC MICROORGANISMS FOR THE RECYCLING OF CARBON FROM
CARBON DIOXIDE AND OTHER INORGANIC CARBON SOURCES THROUGH
CHEMOSYNTHESIS INTO BIOFUEL AND ADDITIONAL USEFUL PRODUCTS." Each
of these applications is incorporated herein by reference in its
entirety for all purposes.
Claims
What is claimed is:
1. A biological and chemical method for the capture and conversion
of an inorganic carbon compound and/or an organic compound
containing only one carbon atom into biomass and/or biochemicals,
comprising: introducing a carbon source consisting of an inorganic
carbon compound and/or an organic compound containing only one
carbon atom into a bioreactor comprising an environment that
comprises a liquid culture medium suitable for maintaining
oxyhydrogen microorganisms and/or capable of maintaining extracts
of oxyhydrogen microorganisms, wherein the liquid culture medium
comprises nitrogen and phosphorous; and converting the inorganic
carbon compound and/or the organic compound containing only one
carbon atom into biomass and/or biochemicals within the environment
via at least one chemosynthetic carbon-fixing reaction utilizing
oxyhydrogen microorganisms belonging to the genus Cupriavidus
and/or cell extracts containing enzymes from the oxyhydrogen
microorganisms belonging to the genus Cupriavidus; wherein the
chemosynthetic fixing reaction is at least partially driven by
chemical and/or electrochemical energy provided by electron donors
comprising gaseous H.sub.2 and electron acceptors comprising
gaseous O.sub.2 that have been generated chemically and/or
electrochemically and/or thermochemically and/or are introduced
into the environment from at least one source external to the
environment, wherein gas phase mixtures of said gaseous H.sub.2 and
O.sub.2, with the concentration of H.sub.2 falling between 4% and
74.5%, are avoided within the bioreactor environment, wherein
O.sub.2 gas is introduced into a first column within the
environment by sparging, bubbling, and/or diffusion of said O.sub.2
gas, and H.sub.2 gas or syngas is introduced into a second column
by sparging, bubbling, and/or diffusion of said H.sub.2 gas or
syngas, wherein there is a liquid connection between the first and
second columns comprising the liquid culture medium, and wherein
the liquid culture medium circulates between the first and second
columns, wherein the biomass and/or biochemicals are produced by
the at least one chemosynthetic carbon-fixing reaction, wherein the
carbon-fixing reaction is maintained using a continuous influx and
removal of nutrient medium and biomass, and wherein concentrations
of the electron donors and the electron acceptors, and levels of
nitrogen and phosphorous, are targeted at constant levels over time
in a steady state, maintained at levels for maximum uptake and
fixation of the inorganic carbon compound and/or the organic
compound containing only one carbon atom and/or for maximum
production of organic compounds comprising amino acids, peptides,
and proteins, wherein surplus biomass is removed from the
bioreactor in order to maintain a constant population and cell
density of said oxyhydrogen microorganisms in the liquid culture
medium, and wherein the biomass and/or biochemicals are separated
from the environment and are processed into a product comprising an
animal feed, a fertilizer, a soil additive, a soil stabilizer, a
carbon source for large scale fermentations, and/or a nutrient
source for the growth of other microbes or organisms.
2. The method according to claim 1, wherein said electron donors
and/or organic compound containing only one carbon atom are
generated through the gasification and/or pyrolysis of organic
matter or through methane steam reforming and provided as a syngas
to the oxyhydrogen microorganisms.
3. The method according to claim 2, wherein the ratio of hydrogen
to carbon monoxide in the syngas is adjusted via the water gas
shift reaction prior to delivery of the syngas to the oxyhydrogen
microorganisms.
4. The method according to claim 1, wherein said thermochemical
and/or electrochemical generation of electron donors and/or
electron acceptors is powered by a carbon dioxide emission-free or
low-carbon emission and/or a renewable source of power, and/or have
been generated by gasification, pyrolysis, or steam reforming of a
waste or biomass feedstock or biogas.
5. The method according to claim 4, wherein the carbon dioxide
emission-free or low-carbon dioxide emission and/or renewable
source of power is utilized for the electrochemical production of
the electron donor.
6. The method according to claim 5, wherein the electrochemical
production of the electron donor comprises electrolysis.
7. The method according to claim 1, wherein said electron donors
and/or electron acceptors are generated or recycled using
renewable, alternative, or conventional sources of power that are
low in greenhouse gas emissions, wherein said source(s) of power is
selected from at least one of: photovoltaics; solar thermal; wind
power; hydroelectric; nuclear; geothermal; enhanced geothermal;
ocean thermal; ocean wave power; and tidal power.
8. The method according to claim 1, wherein said thermochemical
and/or electrochemical generation of electron donors and/or
electron acceptors comprise one or more of the following:
electrolysis of water by approaches including one or more of Proton
Exchange Membranes (PEM), liquid electrolytes, high-pressure
electrolysis, high temperature electrolysis of steam (HTES);
thermochemical splitting of water through one or more of the iron
oxide cycle, cerium(IV) oxide-cerium(III) oxide cycle, zinc
zinc-oxide cycle, sulfur-iodine cycle, copper-chlorine cycle,
calcium-bromine-iron cycle, hybrid sulfur cycle; electrolysis of
hydrogen sulfide; thermochemical splitting of hydrogen sulfide;
other electrochemical or thermochemical processes known to produce
hydrogen with low--or no--carbon dioxide emissions; the
Kvxrner-process and other processes generating a carbon-black
product; carbon capture and sequestration enabled gasification or
pyrolysis of biomass.
9. The method according to claim 1, wherein said electron donors
are generated from pollutants or waste products selected from one
or more of the following: process gas; tail gas; enhanced oil
recovery vent gas; biogas; acid mine drainage; landfill leachate;
landfill gas; geothermal gas; geothermal sludge or brine; metal
contaminants; gangue; tailings; sulfides; disulfides; mercaptans
selected from one or more of methyl and dimethyl mercaptan and
ethyl mercaptan; carbonyl sulfide; carbon disulfide;
alkanesulfonates; dialkyl sulfides; thiosulfate; thiofurans;
thiocyanates; isothiocyanates; thioureas; thiols; thiophenols;
thioethers; thiophene; dibenzothiophene; tetrathionate; dithionite;
thionate; dialkyl disulfides; sulfones; sulfoxides; sulfolanes;
sulfonic acid; dimethyl sulfoniopropionate; sulfonic esters;
hydrogen sulfide; sulfate esters; organic sulfur; sulfur dioxide
and all other sour gases.
10. The method according to claim 1, wherein the oxyhydrogen
microorganisms comprise Cupriavidus necator DSM 531.
11. The method according to claim 1, wherein the inorganic carbon
compound comprises carbon dioxide.
12. The method according to claim 11, wherein the carbon dioxide
comprises carbon dioxide gas, either alone and/or dissolved in a
mixture or solution further comprising carbonate ion and/or
bicarbonate ion.
13. The method according to claim 1, wherein the inorganic carbon
compound comprises carbon contained in a solid phase.
14. The method according to claim 1, wherein the organic compound
containing only one carbon atom comprises carbon monoxide, methane,
methanol, formate, and/or formic acid.
15. The method according to claim 1, wherein said electron donors
further comprise one or more of: ammonia; ammonium; carbon
monoxide; dithionate; elemental sulfur; hydrocarbons;
metabisulfites; nitric oxide; nitrites; sulfates; thiosulfates;
sulfides; sulfites; thionate; thionite; transition metals or their
sulfides; oxides; chalcogenides; halides; hydroxides;
oxyhydroxides; phosphates; sulfates; carbonates, in dissolved or
solid phases; and conduction or valence band electrons in solid
state electrode materials.
16. The method according to claim 1, wherein said electron
acceptors further comprise one or more of the following: carbon
dioxide; nitrites; nitrates; ferric iron or other transition metal
ions; sulfates; or valence or conduction band holes in solid state
electrode materials.
17. The method according to claim 1, wherein the converting step is
preceded by one or more chemical preprocessing steps in which said
electron donors and/or said electron acceptors are generated and/or
refined from at least one input chemical and/or are recycled from
chemicals produced during the chemosynthetic carbon-fixing reaction
and/or chemicals derived from waste streams from other industrial,
mining, agricultural, sewage or waste generating processes.
18. The method according to claim 1, wherein the converting step is
followed by one or more process steps in which any unused nutrients
and/or process water left after removal of oxyhydrogen cell mass
and/or chemical co-products of chemosynthesis and/or waste products
or contaminants of the process stream produced during the fixing
step are recycled back into the environment to support further
chemosynthesis.
19. The method according to claim 1, wherein the carbon fixing
reaction is conducted under aerobic, microaerobic, or facultative
conditions.
20. The method according to claim 1, wherein said biochemicals
comprise an organic chemical product with a carbon chain length of
C5 or greater.
21. The method according to claim 20, wherein the organic chemical
product comprises carbon chain length between C5 and C30.
22. The method according to claim 1, wherein said at least one
chemosynthetic reaction is performed by oxyhydrogen microorganisms
that have been improved, optimized or engineered for the fixation
of the inorganic carbon compound and/or the organic compound
containing only one carbon atom and the production of organic
compounds.
23. The method according to claim 1, wherein the bioreactor does
not comprise transparent materials that expose the oxyhydrogen
microorganisms and/or cell extracts containing enzymes from the
oxyhydrogen microorganisms to light.
24. The method according to claim 1, wherein energy for carbon
fixation is provided by an abiotic process.
25. The method according to claim 1, wherein a feed gas comprising
2% -12% O.sub.2 is introduced into the first column in the
environment.
26. The method according to claim 1, wherein said O.sub.2 gas is
delivered to said bioreactor at a concentration in the range of 2%
to 6%.
27. The method according to claim 1, wherein a feed gas comprising
said H.sub.2 gas and/or syngas is introduced into the environment,
wherein the bioreactor comprises a gas headspace, wherein said
H.sub.2 and/or syngas that is not utilized by the microorganisms in
the chemosynthetic carbon-fixing reaction passes through the liquid
culture medium and into the gas headspace and is recirculated by
pumping the un-utilized gas out of the headspace, compressing it,
and pumping it back into the liquid culture medium.
28. The method according to claim 1, wherein said recirculated
H.sub.2 and/or syngas is pumped back into the liquid culture medium
at the bottom of the second column in said bioreactor.
29. The method according to claim 1, wherein said biomass comprises
a dry biomass concentration of at least 4 g/1 in steady state in
said continuous microbial culture.
30. The method according to claim 1, wherein said O.sub.2 gas in
the culture medium comprises dissolved oxygen that is controlled
and targeted at a constant level over time in a steady state.
31. The method according to claim 1, wherein the bioreactor
comprises a gas headspace, and wherein oxygen concentration in the
headspace is controlled and targeted at a constant level over time
in a steady state.
32. The method according to claim 1, wherein the bioreactor
comprises a gas headspace, and wherein hydrogen concentration in
the headspace is controlled and targeted at a constant level over
time in a steady state.
33. The method according to claim 1, wherein said H.sub.2 gas is
introduced into the second column in the environment by bubbling
through the liquid culture medium and/or by diffusing through a
membrane that contacts the liquid culture medium and that is
impermeable to the liquid culture medium, and wherein said O.sub.2
gas is pumped into the liquid culture medium using sparging
equipment, a diffuser, a bubble aerator, and/or venturi
equipment.
34. The method according to claim 1, wherein the said liquid medium
flows in the direction of the introduced O.sub.2 gas and the liquid
medium flows countercurrent to the direction of the introduced
H.sub.2 gas or syngas.
35. The method according to claim 1, wherein the liquid medium
flows countercurrent to the direction of the introduced O.sub.2 gas
and the liquid medium flows in the direction of the introduced
H.sub.2 gas or syngas.
36. The method according to claim 1, wherein the O.sub.2 gas is
bubbled forcefully such that the first column acts as an airlift
reactor and drives the circulation of the culture medium between
the first and second columns.
37. The method according to claim 1, wherein the circulation of the
liquid medium is assisted by impellers, turbines, and/or pumps.
38. The method according to claim 1, wherein the oxyhydrogen
microorganisms freely circulate along with the liquid medium
between the said first and second columns.
Description
FIELD OF INVENTION
The present invention falls within the technical areas of biofuels,
bioremediation, carbon capture, carbon dioxide-to-fuels, carbon
recycling, carbon sequestration, energy storage, gas-to-liquids,
waste energy to fuels, syngas conversions, and
renewable/alternative and/or low carbon dioxide emission sources of
energy. Specifically the present invention is a unique example of
the use of biocatalysts within a biological and chemical process to
fix carbon dioxide and/or other forms of inorganic carbon and/or or
other C1 carbon sources into longer carbon chain organic chemical
products in a non-photosynthetic process powered by low carbon
emission energy sources and/or waste energy sources. In addition
the present invention involves the production of chemical
co-products that arc co-generated through carbon-fixation reaction
steps and/or non-biological reaction steps as part of an overall
carbon capture and conversion process or syngas conversion process.
The present invention can enable the effective and economic capture
of carbon dioxide from the atmosphere or from a point source of
carbon dioxide emissions as well as the economic use of waste
energy sources and/or renewable energy sources and/or low carbon
emission energy sources, for the production of liquid
transportation fuel and/or other organic chemical products, which
will help address greenhouse gas induced climate change and
contribute to the domestic production of renewable liquid
transportation fuels and/or other organic chemicals without any
dependence upon agriculture.
BACKGROUND
Great interest and resources have been directed towards developing
technologies that use renewable energy or waste energy for the
conversion of carbon dioxide, or other low value carbon sources,
into useful organic chemicals in order to provide alternatives to
chemicals, materials and fuels derived from petroleum or other
fossil sources. Most of the focus in the area of CO.sub.2
conversion has been placed on biological approaches that utilize
photosynthesis to fix CO.sub.2 into biomass or end-products, while
some effort has been directed at fully abiotic and chemical
processes for fixing CO.sub.2.
A type of CO.sub.2-to-organic chemical approach that has received
relatively less attention is hybrid chemical/biological processes
where the biological step is limited to CO.sub.2 fixation alone,
corresponding to the dark reaction of photosynthesis. The potential
advantages of such a hybrid CO.sub.2-to-organic chemical process
include the ability to combine enzymatic capabilities gained
through billions of years of evolution in fixing CO.sub.2, with a
wide array of abiotic technologies to power the process such as
solar PV, solar thermal, wind, geothermal, hydroelectric, or
nuclear. Microorganisms performing carbon fixation without light
can be contained in more controlled and protected environments,
less prone to water and nutrient loss, contamination, or weather
damage, than what can be used for culturing photosynthetic
microorganisms. Furthermore an increase in bioreactor capacity can
be met with vertical rather than horizontal construction, making it
potentially far more land efficient. A hybrid chemical/biological
system offers the possibility of a CO.sub.2-to-organic chemical
process that avoids many drawbacks of photosynthesis while
retaining the biological capabilities for complex organic synthesis
from CO.sub.2.
Chemoautotrophic microorganisms are generally microbes that can
perform CO.sub.2 fixation like in the photosynthetic dark reaction,
but which can get the reducing equivalents needed for CO.sub.2
fixation from an inorganic external source, rather than having to
internally generate them through the photosynthetic light reaction.
Carbon fixing biochemical pathways that occur in chemoautotrophs
include the reductive tricarboxylic acid cycle, the
Calvin-Benson-Bassham cycle, and the Wood-Ljungdahl pathway.
Prior work is known relating to certain applications of
chemoautotrophic microorganisms in the capture and conversion of
CO.sub.2 gas to fixed carbon. However, many of these approaches
have suffered shortcomings that have limited the effectiveness,
economic feasibility, practicality and commercial adoption of the
described processes. The present invention in certain aspects
addresses one or more of the aforementioned shortcomings.
It is believed that the present invention utilizing oxyhydrogen
microorganisms in the chemosynthetic fixation of CO.sub.2 under
carefully controlled oxygen levels may have advantages for the
production of longer chain organic compounds (e.g., C.sub.5 and
longer). The ability to produce longer chain organic compounds is
an important advantage for the present invention since the energy
densities (energy per unit volume) are generally higher for longer
chain organic compounds, and the compatibility with the current
transportation fleet is generally greater relative to, for example,
shorter chain products such as C1 and C2 products.
SUMMARY OF THE INVENTION
In response to a need in the art that the inventors have recognized
in making the invention, a novel combined biological and chemical
process for the capture and conversion of inorganic carbon and/or
C1 carbon sources to longer chain organic compounds, and
particularly organic compounds with C5 or longer chain lengths,
through the use of oxyhydrogen microorganisms for carbon capture
and fixation is described. In some embodiments, the process can
couple the efficient production of high value organic compounds
such as liquid hydrocarbon fuel with the disposal of waste sources
of carbon, as well as with the capture of CO.sub.2, which can
generate additional revenue.
In one aspect, a biological and chemical method for the capture and
conversion of an inorganic carbon compound and/or an organic
compound containing only one carbon atom into an organic chemical
product is described. In some embodiments, the method comprises
introducing an inorganic carbon compound and/or an organic compound
containing only one carbon atom into an environment suitable for
maintaining oxyhydrogen microorganisms and/or capable of
maintaining extracts of oxyhydrogen microorganisms; and converting
the inorganic carbon compound and/or the organic compound
containing only one carbon atom into the organic chemical product
and/or a precursor thereof within the environment via at least one
chemosynthetic carbon-fixing reaction utilizing the oxyhydrogen
microorganisms and/or cell extracts containing enzymes from the
oxyhydrogen microorganisms. In some embodiments, the chemosynthetic
fixing reaction is at least partially driven by chemical and/or
electrochemical energy provided by electron donors and electron
acceptors that have been generated chemically and/or
electrochemically and/or are introduced into the environment from
at least one source external to the environment.
In one aspect, a bioreactor is described. The bioreactor comprises,
in one set of embodiments, a first column comprising an upper
portion and a lower portion; and a second column comprising an
upper portion and a lower portion, the upper portion of the second
column fluidically connected to the upper portion of the first
column, and the lower portion of the second column fluidically
connected to the lower portion of the first column. In some
embodiments, the bioreactor is constructed and arranged such that,
when a liquid is circulated between the first and second columns, a
volume of gas is substantially stationary at the top of the first
column and/or the second column. In some embodiments, the volume of
gas occupies at least about 2% of the total volume of the column in
which the volume is positioned.
In another aspect, a method of operating a bioreactor is provided.
The method comprises, in some embodiments, circulating a liquid
comprising a growth medium between a first column and a second
column, wherein, during operation, a volume of gas remains
substantially stationary at the top of the first column and/or the
second column, and the volume of gas occupies at least about 2% of
the total volume of the column in which the volume is
positioned.
In one aspect, an electrolysis device is provided. In some
embodiments, the electrolysis device comprises a chamber
constructed and arranged to electrolyze water to produce oxygen and
hydrogen; and an outlet comprising a separator constructed and
arranged to separate at least a portion of the oxygen within a
stream from at least a portion of the hydrogen within a stream such
that the hydrogen content of the fluid exiting the separator is
suitable for use as a feed stream to a reactor containing a culture
of oxyhydrogen microorganisms.
In another aspect, a method of operating an electrolysis device is
described. The method comprises, in some embodiments, electrolyzing
water to produce a first stream containing oxygen and hydrogen; and
separating at least a portion of the oxygen from at least a portion
of the hydrogen to produce a second stream relatively rich in
hydrogen compared to the first stream, wherein the second stream is
suitable for use as a feed stream to a reactor containing a culture
of oxyhydrogen microorganisms.
The present invention, in certain embodiments, provides
compositions and methods for the capture of carbon dioxide from
carbon dioxide-containing gas streams and/or atmospheric carbon
dioxide or carbon dioxide in dissolved, liquefied or
chemically-bound form through a chemical and biological process
that utilizes obligate or facultative oxyhydrogen microorganisms,
and/or cell extracts containing enzymes from oxyhydrogen
microorganisms in one or more carbon fixing process steps.
The present invention, in certain embodiments, provides
compositions and methods for the utilization of C1 carbon sources
including but not limited to carbon monoxide, methane, methanol,
formate, or formic acid, and/or mixtures containing C1 chemicals
including but not limited to various syngas compositions generated
from various gasified, pyrolyzed, or steam-reformed fixed carbon
feedstocks, and convert said C1 chemicals into longer chain organic
compounds,
The present invention, in certain embodiments, provides
compositions and methods for the recovery, processing, and use of
the organic compounds produced by chemosynthetic reactions
performed by oxyhydrogen microorganisms to fix inorganic carbon
and/or C1 carbon sources into longer chain organic compounds. The
present invention, in certain embodiments, provides compositions
and methods for the maintenance and control of the oxygen levels in
the carbon-fixation environment for the enhanced (e.g., optimal)
production of C5 or longer organic compound products through carbon
fixation. The present invention, in certain embodiments, provides
compositions and methods for the generation, processing and
delivery of chemical nutrients needed for carbon-fixation and
maintenance of oxyhydrogen microorganism cultures, including but
not limited to the provision of electron donors and electron
acceptors needed for non-photosynthetic carbon-fixation. The
present invention, in certain embodiments, provides compositions
and methods for the maintenance of an environment conducive for
carbon-fixation, and the recovery and recycling of unused chemical
nutrients and process water.
The present invention, in certain embodiments, provides
compositions and methods for chemical process steps that occur in
series and/or in parallel with the chemosynthetic reaction steps
that: convert unrefined raw input chemicals to more refined
chemicals that are suited for supporting the chemosynthetic carbon
fixing step; that convert energy inputs into a chemical form that
can be used to drive chemosynthesis, and specifically into chemical
energy in the form of electron donors and electron acceptors; that
direct inorganic carbon captured from industrial or atmospheric or
aquatic sources to the carbon fixation steps of the process under
conditions that are suitable to support chemosynthetic carbon
fixation by the oxyhydrogen microorganisms or enzymes and/or direct
C1 chemicals derived from low value or waste sources of carbon such
as carbon monoxide, methane, methanol, formate, or formic acid,
and/or mixtures containing C1 chemicals including but not limited
to various syngas compositions derived from the gasification,
pyrolysis, or steam reforming of various low value or waste carbon
sources, that can be used by the oxyhydrogen microorganism as a
carbon sources and any energy source for the synthesis of longer
chain organic chemicals; that further process the output products
of the carbon fixation steps into a form suitable for storage,
shipping, and sale, and/or safe disposal in a manner that results
in a net reduction of gaseous CO.sub.2 released into the atmosphere
and/or the upgrade of a low value or waste material into a finished
chemical, fuel, or nutritional product. The fully chemical process
steps combined with the chemosynthetic carbon fixation steps
constitute the overall carbon capture and conversion process of
some embodiments of the present invention.
One feature of certain embodiments of the present invention is the
inclusion of one or more process steps within a chemical process
for the capture of inorganic carbon and conversion to fixed carbon
products, that utilize oxyhydrogen microorganisms and/or enzymes
from oxyhydrogen microorganisms as a biocatalyst for the fixation
of carbon dioxide in carbon dioxide-containing gas streams or the
atmosphere or water and/or dissolved or solid forms of inorganic
carbon, into organic compounds. In some such embodiments carbon
dioxide containing flue gas, or process gas, or air, or inorganic
carbon in solution as dissolved carbon dioxide, carbonate ion, or
bicarbonate ion including aqueous solutions such as sea water, or
inorganic carbon in solid phases such as but not limited to
carbonates and bicarbonates, is pumped or otherwise added to a
vessel or enclosure containing nutrient media and oxyhydrogen
microorganisms. In some such cases oxyhydrogen microorganisms
perform chemosynthesis to fix inorganic carbon into organic
compounds using the chemical energy stored in molecular hydrogen
and/or valence or conduction electrons in solid state electrode
materials and/or one or more of the following list of electron
donors pumped or otherwise provided to the nutrient media including
but not limited to: ammonia; ammonium; carbon monoxide; dithionite;
elemental sulfur; hydrocarbons; metabisulfites; nitric oxide;
nitrites; sulfates such as thiosulfates including but not limited
to sodium thiosulfate (Na.sub.2S.sub.2O.sub.3) or calcium
thiosulfate (CaS.sub.2O.sub.3); sulfides such as hydrogen sulfide;
sulfites; thionate; thionite; transition metals or their sulfides,
oxides, chalcogenides, halides, hydroxides, oxyhydroxides,
phosphates, sulfates, or carbonates, in soluble or solid phases. In
some embodiments, conduction or valence band electrons in solid
state electrode materials can be used. The electron donors can be
oxidized by electron acceptors in the chemosynthetic reaction.
Electron acceptors that may be used at the chemosynthetic reaction
step include oxygen and/or other electron acceptors including but
not limited to one or more of the following: carbon dioxide, ferric
iron or other transition metal ions, nitrates, nitrites, sulfates,
oxygen, or valence or conduction band holes in solid state
electrode materials.
One feature of certain embodiments of the present invention is the
inclusion of one or more process steps within a chemical process
for the conversion of C1 carbon sources including but not limited
to carbon monoxide, methane, methanol, formate, or formic acid,
and/or mixtures containing C1 chemicals including but not limited
to various syngas compositions generated from various gasified,
pyrolyzed, or steam-reformed fixed carbon feedstocks, that utilize
oxyhydrogen microorganisms and/or enzymes from oxyhydrogen
microorganisms as a biocatalyst for the conversion of C1 chemicals
into longer chain organic chemicals (i.e. C2 or longer and, in some
embodiments, C5 or longer carbon chain molecules). In some such
embodiments C1 containing syngas, or process gas, or C1 chemicals
in a pure liquid form or dissolved in solution is pumped or
otherwise added to a vessel or enclosure containing nutrient media
and oxyhydrogen microorganisms. In some such cases oxyhydrogen
microorganisms perform biochemical synthesis to elongate C1
chemicals into longer carbon chain organic chemicals using the
chemical energy stored in the C1 chemical, and/or molecular
hydrogen and/or valence or conduction electrons in solid state
electrode materials and/or one or more of the following list of
electron donors pumped or otherwise provided to the nutrient media
including but not limited to: ammonia; ammonium; carbon monoxide;
dithionite; elemental sulfur; hydrocarbons; metabisulfites; nitric
oxide; nitrites; sulfates such as thiosulfates including but not
limited to sodium thiosulfate (Na.sub.2S.sub.2O.sub.3) or calcium
thiosulfate (CaS.sub.2O.sub.3); sulfides such as hydrogen sulfide;
sulfites; thionate; thionite; transition metals or their sulfides,
oxides, chalcogenides, halides, hydroxides, oxyhydroxides,
sulfates, or carbonates, in soluble or solid phases. The electron
donors can be oxidized by electron acceptors in a chemosynthetic
reaction. Electron acceptors that may be used at this reaction step
include oxygen and/or other electron acceptors including but not
limited to one or more of the following: carbon dioxide, ferric
iron or other transition metal ions, nitrates, nitrites, oxygen, or
holes in solid state electrode materials.
The chemosynthetic reaction step or steps of the process whereby
carbon dioxide and/or inorganic carbon is fixed into organic carbon
in the form of organic compounds and biomass and/or the reaction
steps converting C1 chemicals to longer chain organic chemicals
whereby a C1 chemical such as but not limited to carbon monoxide,
methane, methanol, formate, or formic acid, and/or mixtures
containing C1 chemicals including but not limited to various syngas
compositions generated from various gasified, pyrolyzed, or
steam-reformed fixed carbon feedstocks, are biochemically converted
into longer chain organic chemicals (i.e. C2 or longer and, in some
embodiments, C5 or longer carbon chain molecules) can be performed
in aerobic, microaerobic, anoxic, anaerobic conditions, or
facultative conditions. A facultative environment is considered to
be one having aerobic upper layers and anaerobic lower layers
caused by stratification of the water column.
The oxygen level is controlled in some embodiments of the current
invention so that the production of targeted organic compounds by
the oxyhydrogen microorganisms through carbon-fixation is
controlled (e.g., optimized). One objective of controlling oxygen
levels is to control (e.g., optimize) the intracellular Adenosine
Triphosphate (ATP) concentration through the cellular reduction of
oxygen and production of ATP by oxidative phosphorylation, while
simultaneously keeping the environment sufficiently reducing so
that a high ratio of NADH (or NADPH) to NAD (or NADP) is also
maintained.
An advantage of using oxyhydrogen microorganisms over strictly
anaerobic acetogenic or methanogenic microorganisms for carbon
capture applications and/or syngas conversion applications is the
higher oxygen tolerance of oxyhydrogen microorganisms.
A further advantage of using oxyhydrogen microorganisms for carbon
capture applications and/or syngas conversion applications and/or
biofuel production over using acetogens is that the production of
ATP powered by the oxyhydrogen reaction results in a water product,
which can readily be incorporated into the process stream, rather
than the generally undesirable acetic acid or butyric acid products
of acidogenesis which can harm the microorganisms by dropping the
solution pII or accumulating to toxic levels.
An additional feature of certain embodiments of the present
invention regards the source, production, or recycling of the
electron donors used by the oxyhydrogen microorganisms to fix
carbon dioxide into organic compounds and/or to synthesize longer
carbon chain organic molecules from C1 chemicals. The electron
donors used for carbon dioxide capture and carbon fixation can be
produced or recycled in certain embodiments of the present
invention electrochemically or thermochemically using power from a
number of different renewable and/or low carbon emission energy
technologies including but not limited to: photovoltaics, solar
thermal, wind power, hydroelectric, nuclear, geothermal, enhanced
geothermal, ocean thermal, ocean wave power, tidal power. The
electron donors can also be of mineralogical origin including but
not limited to reduced S and Fe containing minerals. The electron
donors used in certain embodiments of the present invention can
also be produced or recycled through chemical reactions with
hydrocarbons that may or may not be a non-renewable fossil fuel,
but where said chemical reactions produce low or zero carbon
dioxide gas emissions. For example oxide reduction reactions that
produce a carbonate and a hydrogen product that can be used as
electron donor in the carbon-fixation reaction steps of certain
embodiments of the present invention include:
2CH.sub.4+Fe.sub.2O.sub.3+3H.sub.2O2FeCO.sub.3+7H.sub.2 and/or
CH.sub.4+CaO+2H.sub.2OCaCO.sub.3+4H.sub.2.
An additional feature of certain embodiments of the present
invention regards the formation and recovery of organic compounds
and/or biomass from the chemosynthetic carbon fixation step or
steps. These organic compounds and/or biomass products can have a
variety of applications.
An additional feature of certain embodiments of the present
invention regards using modified oxyhydrogen microorganisms in the
carbon-fixation step/steps such that a superior quantity and/or
quality of organic compounds, biochemicals, or biomass is generated
through chemosynthesis. The oxyhydrogen microbes used in these
steps may be modified through artificial means including but not
limited to accelerated mutagenesis (e.g. using ultraviolet light or
chemical treatments), genetic engineering or modification,
hybridization, synthetic biology or traditional selective breeding.
Possible modifications of the oxyhydrogen microorganisms include
but are not limited to those directed at producing increased
quantity and/or quality of organic compounds and/or biomass to be
used as a biofuels, or as feedstock for the production of biofuels
including, but not limited to JP-8 jet fuel, diesel, gasoline,
biodiesel, butanol, ethanol, hydrocarbons, methane, and
pseudovegetable oil or any other hydrocarbon suitable for use as a
renewable/alternate fuel leading to lowered greenhouse gas
emissions.
Also described are compositions and methods that reduce the hazards
of performing gas fermentations that utilize mixtures of hydrogen
and oxygen within the invented process.
Compositions and methods that take advantage of the oxygen
tolerance and ability to use oxygen as an electron acceptor
possessed by oxyhydrogen microorganisms in order to enable a system
for converting water into hydrogen or hydride electron donors and
oxygen electron acceptors, that has improved efficiency over the
application of current state-of-the-art electrolysis for the
purpose of generating hydrogen or hydride electron donors and
oxygen electron acceptors, are also described.
Also described are process steps for the recovery and further
finishing of useful chemicals produced both by the biological
carbon fixation steps of the process, as well as from
non-biological process steps.
Other advantages and novel features of the present invention will
become apparent from the following detailed description of various
non-limiting embodiments of the invention when considered in
conjunction with the accompanying figures. All publications, patent
applications and patents mentioned in the text are incorporated by
reference in their entirety. In cases where the present
specification and a document incorporated by reference include
conflicting and/or inconsistent disclosure, the present
specification shall control.
BRIEF DESCRIPTION OF THE DRAWINGS
Non-limiting embodiments of the present invention will be described
by way of example with reference to the accompanying figures, which
are schematic and are not intended to be drawn to scale. For
purposes of clarity, not every component is labeled in every
figure, nor is every component of each embodiment of the invention
shown where illustration is not necessary to allow those of
ordinary skill in the art to understand the invention. In the
figures:
FIG. 1 is a general process flow diagram for one embodiment of this
invention for a carbon capture and fixation process;
FIG. 2 is a process flow diagram for another embodiment of the
present invention with capture of CO.sub.2 performed by a
microorganism capable of performing an oxyhydrogen reaction (e.g.,
hydrogen oxidizing purple non-sulfur bacteria) to produce a
lipid-rich biomass that is converted into JP-8 jet fuel;
FIG. 3 is diagram of a bioreactor design that can avoid dangerous
mixtures of hydrogen and oxygen by exploiting the low solubilities
of hydrogen and oxygen gas in water while providing the oxyhydrogen
microorganism with the oxygen and hydrogen needed for cellular
energy and carbon fixation;
FIG. 4 is a diagram of a bioreactor design that takes advantage of
the relatively high solubility of carbon dioxide and the strong
ability of oxyhydrogen microorganism to capture carbon dioxide from
relatively dilute streams using a carbon concentrating mechanism
(CCM), to remove CO.sub.2 from a dilute gas mixture and separate it
from low solubility gases such as oxygen and nitrogen; and
FIG. 5 is an electrolysis technology that is specially designed to
take advantage of the oxyhydrogen microorganisms' tolerance and
need for a certain concentration of oxygen by decreasing the
complete separation of the hydrogen and oxygen produced from
standard electrolysis.
DETAILED DESCRIPTION
The present invention provides, in certain embodiments,
compositions and methods for the capture and fixation of carbon
dioxide from carbon dioxide-containing gas streams and/or
atmospheric carbon dioxide or carbon dioxide in liquefied or
chemically-bound form through a chemical and biological process
that utilizes obligate or facultative oxyhydrogen microorganisms,
and/or cell extracts containing enzymes from oxyhydrogen
microorganisms in one or more process steps. The fixation of
inorganic carbon sources other than CO.sub.2 and/or other C1 carbon
sources are also described. Cell extracts include but are not
limited to: a lysate, extract, fraction or purified product
exhibiting chemosynthetic enzyme activity that can be created by
standard methods from oxyhydrogen microorganisms. In addition the
present invention, in certain embodiments, provides compositions
and methods for the recovery, processing, and use of the chemical
products of chemosynthetic reaction step or steps performed by
oxyhydrogen microorganisms to fix inorganic carbon into organic
compounds and/or synthetic reaction step or steps performed by
oxyhydrogen microorganisms to elongate C1 molecules to longer
carbon chain organic chemicals. Finally the present invention, in
certain embodiments, provides compositions and methods for the
production and processing and delivery of chemical nutrients needed
for chemoautotrophic carbon-fixation by the oxyhydrogen
microorganisms, and particularly electron donors including but not
limited to molecular hydrogen and/or electrical power, and electron
acceptors including but not limited to oxygen and carbon dioxide to
drive the carbon fixation reaction; compositions and methods for
the maintenance of an environment conducive for carbon-fixation by
oxyhydrogen microorganisms; and compositions and methods for the
removal of the chemical products of chemosynthesis from the
oxyhydrogen culture environment and the recovery and recycling of
unused of chemical nutrients.
The terms "molecular hydrogen," "dihydrogen," and "II.sub.2" are
used interchangeably throughout.
The terms "oxyhydrogen microorganism" and "knallgas microorganism"
are used interchangeably throughout. Oxyhydrogen microorganisms are
generally described in Chapter 5, Section III of Thermophilic
Bacteria, a book by Jakob Kristjansson, CRC Press, 1992, which is
incorporated herein by reference. Generally, oxyhydrogen
microorganisms are capable of performing the oxyhydrogen reaction.
Oxyhydrogen microorganisms generally have the ability to use
molecular hydrogen by means of hydrogenases with some of the
electrons donated from H.sub.2 being utilized for the reduction of
NAD.sup.+ (and/or other intracellular reducing equivalents) and the
rest of the electrons for aerobic respiration. In addition,
oxyhydrogen microorganisms generally are capable of fixing CO.sub.2
autotrophically, through pathways such as the reverse Calvin Cycle
or the reverse citric acid cycle.
In addition, the terms "oxyhydrogen reaction" and "knallgas
reaction" are used interchangeably throughout to refer to the
microbial oxidation of molecular hydrogen by molecular oxygen. The
oxyhydrogen reaction is generally expressed as:
2H.sub.2+O.sub.22H.sub.2O+energy and/or by stoichiometric
equivalents of this reaction.
Exemplary oxyhydrogen microorganisms that can be used in one or
more process steps of certain embodiments of the present invention
include but are not limited to one or more of the following: purple
non-sulfur photosynthetic bacteria including but not limited to
Rhodopseudomonas palustris, Rhodopseudomonas capsulata,
Rhodopseudomonas viridis, Rhodopseudomonas sulfoviridis,
Rhodopseudomonas blastica, Rhodopseudomonas spheroides,
Rhodopseudomonas acidophila and other Rhodopseudomonas sp.,
Rhodospirillum rubrum, and other Rhodospirillum sp.; Rhodococcus
opacus and other Rhodococcus sp.; Rhizobium japonicum and other
Rhizobium sp.; Thiocapsa roseopersicina and other Thiocapsa sp.;
Pseudomonas hydrogenovora, Pseudomonas hydrogenothermophila, and
other Pseudomonas sp.; Hydrogenomonas pantotropha, Hydrogenomonas
eutropha, Hydrogenomonas facilis, and other Hydrogenomonas sp.;
Hydrogenobacter thermophilus and other Hydrogenobacter sp.;
Hydrogenovibrio marinus and other Hydrogenovibrio sp.; Helicobacter
pylori and other Helicobacter sp.; Xanthobacter sp.; Hydrogenophaga
sp.; Bradyrhizobium japonicum and other Bradyrhizobium sp.;
Ralstonia eutropha and other Ralstonia sp.; Alcaligenes eutrophus
and other Alcaligenes sp.; Variovorax paradoxus, and other
Variovorax sp.; Acidovorax facilis, and other Acidovorax sp.;
cyanobacteria including but not limited to Anabaena oscillarioides,
Anabaena spiroides, Anabaena cylindrica, and other Anabaena sp.;
green algae including but not limited to Scenedesmus obliquus and
other Scene desmus sp., Chlamydomonas reinhardii and other
Chlamydomonas sp., Ankistrodesmus sp., Rhaphidium polymorphium and
other Rhaphidium sp.; as well as a consortiums of microorganisms
that include oxyhydrogen microorganisms.
The different oxyhydrogen microorganisms that can be used in
certain embodiments of the present invention may be native to a
range environments including but not limited to hydrothermal vents,
geothermal vents, hot springs, cold seeps, underground aquifers,
salt lakes, saline formations, mines, acid mine drainage, mine
tailings, oil wells, refinery wastewater, oil, gas, or hydrocarbon
contaminated waters; coal seams, the deep sub-surface, waste water
and sewage treatment plants, geothermal power plants, sulfatara
fields, soils including but not limited to soils contaminated with
hydrocarbons and/or located under or around oil or gas wells, oil
refineries, oil pipelines, gasoline service stations. They may or
may not be extremophiles including but not limited to thermophiles,
hyperthermophiles, acidophiles, halophiles, and psychrophiles.
In some embodiments, relatively long-chain chemical products can be
produced. For example, the organic chemical product produced in
some embodiments can include compounds with carbon chain lengths of
at least C5, at least C10, at least C15, at least C20, between
about C5 and about C30, between about C10 and about C30, between
about C15 and about C30, or between about C20 and about C30.
FIG. 1 illustrates the general process flow diagram for embodiments
of the present invention that have a process step for the
generation of electron donors (e.g., molecular hydrogen electron
donors) suitable for supporting chemosynthesis from an energy input
and raw inorganic chemical input; followed by recovery of chemical
co-products from the electron donor generation step; delivery of
generated electron donors along with oxygen electron acceptors,
water, nutrients, and CO.sub.2 from a point industrial flue gas
source, into chemosynthetic reaction step or steps that make use of
oxyhydrogen microorganisms to capture and fix carbon dioxide,
creating chemical and biomass co-products through chemosynthetic
reactions; followed by process steps for the recovery of both
chemical and biomass products from the process stream; and
recycling of unused nutrients and process water, as well as cell
mass needed to maintain the microbial culture, back into the
carbon-fixation reaction steps.
In the embodiment illustrated in FIG. 1, the CO.sub.2 containing
flue gas is captured from a point source or emitter. Electron
donors (e.g., H.sub.2) needed for chemosynthesis can be generated
from input inorganic chemicals and energy. The flue gas can be
pumped through bioreactors containing oxyhydrogen microorganisms
along with electron donors and acceptors needed to drive
chemosynthesis and a medium suitable to support the microbial
culture and carbon fixation through chemosynthesis. The cell
culture may be continuously flowed into and out of the bioreactors.
After the cell culture leaves the bioreactors, the cell mass can be
separated from the liquid medium. Cell mass needed to replenish the
cell culture population at a desirable (e.g., optimal) level can be
recycled back into the bioreactor. Surplus cell mass can be dried
to form a dry biomass product which can be further post-processed
into various chemical, fuel, or nutritional products. Following the
cell separation step, extracellular chemical products of the
chemosynthetic reaction can be removed from the process flow and
recovered. Then, any undesirable waste products that might be
present are removed. Following this, the liquid medium and any
unused nutrients can be recycled back into the bioreactors.
Many of the reduced inorganic chemicals upon which chemoautotrophs
grow (e.g. H.sub.2, H.sub.2S, ferrous iron, ammonium, Mn.sup.2+)
can be readily produced using electrochemical and/or thermochemical
processes known in the art of chemical engineering that may
optionally be powered by a variety carbon dioxide emission-free or
low-carbon emission and/or renewable sources of power including
wind, hydroelectric, nuclear, photovoltaics, or solar thermal.
Certain embodiments of the present invention use carbon dioxide
emission-free or low-carbon emission and/or renewable sources of
power in the production of electron donors including but not
limited to one or more of the following: photovoltaics, solar
thermal, wind power, hydroelectric, nuclear, geothermal, enhanced
geothermal, ocean thermal, ocean wave power, tidal power. In
certain embodiments of the present invention oxyhydrogen
microorganisms function as biocatalysts for the conversion of
renewable energy and/or low or zero carbon emission energy into
liquid hydrocarbon fuel, or high energy density organic compounds
generally, with CO.sub.2 captured from flue gases, or from the
atmosphere, or ocean serving as a carbon source. These embodiments
of the present invention can provide renewable energy technologies
with the capability of producing a transportation fuel having
significantly higher energy density than if the renewable energy
sources are used to produce hydrogen gas--which must be stored in
relatively heavy storage systems (e.g. tanks or storage
materials)--or if it is used to charge batteries, which have
relatively low energy density. Additionally the liquid hydrocarbon
fuel product of certain embodiments of the present invention may be
more compatible with the current transportation infrastructure
compared to battery or hydrogen energy storage options.
The position of the process step or steps for the generation of
electron donors (e.g., molecular hydrogen electron donors) in the
general process flow of certain embodiments of the present
invention is illustrated in FIG. 1 by Box 3, labeled "Electron
Donor Generation." Electron donors produced in certain embodiments
of the present invention using electrochemical and/or
thermochemical processes known in the art of chemical engineering
and/or generated from natural sources include, but are not limited
to molecular hydrogen and/or valence or conduction electrons in
solid state electrode materials and/or other reducing agents
including but are not limited to one or more of the following:
ammonia; ammonium; carbon monoxide; dithionite; elemental sulfur;
hydrocarbons; metabisulfites; nitric oxide; nitrites; sulfates such
as thiosulfates including but not limited to sodium thiosulfate
(Na.sub.2S.sub.2O.sub.3) or calcium thiosulfate (CaS.sub.2O.sub.3);
sulfides such as hydrogen sulfide; sulfites; thionate; thionite;
transition metals or their sulfides, oxides, chalcogenides,
halides, hydroxides, oxyhydroxides, sulfates, or carbonates, in
soluble or solid phases.
Certain embodiments of the present invention use molecular hydrogen
as the electron donor. Hydrogen electron donors are generated by
methods known in to art of chemical and process engineering
including but not limited to one or more of the following: through
electrolysis of water by approaches including but not limited to
using Proton Exchange Membranes (PEM), liquid electrolytes such as
KOH, high-pressure electrolysis, high temperature electrolysis of
steam (HTES); thermochemical splitting of water through methods
including but not limited to the iron oxide cycle, cerium(IV)
oxide-cerium(III) oxide cycle, zinc zinc-oxide cycle, sulfur-iodine
cycle, copper-chlorine cycle, calcium-bromine-iron cycle, hybrid
sulfur cycle; electrolysis of hydrogen sulfide; thermochemical
and/or electrochemical splitting of hydrogen sulfide; other
electrochemical or thermochemical processes known to produce
hydrogen with low- or no-carbon dioxide emissions including but not
limited to: carbon capture and sequestration enabled methane
reforming; carbon capture and sequestration enabled coal
gasification; the Kv.ae butted.rner-process and other processes
generating a carbon-black product; carbon capture and sequestration
enabled gasification or pyrolysis of biomass; and the half-cell
reduction of H.sup.+ to H.sub.2 accompanied by the half-cell
oxidization of electron sources including but not limited to
ferrous iron (Fe.sup.2+) oxidized to ferric iron (Fe.sup.3+) or the
oxidation of sulfur compounds whereby the oxidized iron or sulfur
can be recycled to back to a reduced state through additional
chemical reaction with minerals including but not limited to metal
sulfides, hydrogen sulfide, or hydrocarbons.
In certain embodiment of the present invention, hydrogen electron
donors are not necessarily generated with low- or no-carbon dioxide
emissions, however the hydrogen is generated from waste or low
value sources of energy using methods known in to art of chemical
and process engineering including but not limited to gasification,
pyrolysis, or steam-reforming of feedstock such as but not limited
to municipal waste, black liquor, agricultural waste, wood waste,
stranded natural gas, biogas, sour gas, methane hydrates, tires,
sewage, manure, straw, and low value, highly lignocellulosic
biomass in general.
In certain embodiments of the present invention that utilize
molecular hydrogen as an electron donor for the carbon-fixation
reactions performed by oxyhydrogen microorganisms, there can be a
chemical co-product formed in the generation of molecular hydrogen
using a renewable and/or CO.sub.2 emission-free energy input. If
water is used as a hydrogen source, then oxygen can be a co-product
of water splitting through processes including but not limited to
electrolysis or thermochemical water splitting. In certain
embodiments of the present invention using water as a hydrogen
source, some of the oxygen co-product can be used in the
oxyhydrogen carbon fixation step for the production of
intracellular ATP through the oxyhydrogen reaction enzymatically
linked to oxidative phosphorylation. In certain embodiments of the
present invention, the oxygen produced by water-splitting in excess
of what is required to maintain favorable (e.g., optimal)
conditions for carbon fixation and organic compound production by
the oxyhydrogen microorganisms can be processed into a form
suitable for sale through process steps known in the art and
science of commercial oxygen gas production. In certain embodiments
of the present invention where hydrogen sulfide is the hydrogen
source, sulfur or sulfuric acid can be a chemical co-product of
molecular hydrogen production. In certain embodiments of the
present invention where sulfuric acid is a co-product of hydrogen
production, some of the sulfuric acid can be used in the hydrolysis
of biomass in post-carbon fixation process steps. In certain
embodiments of the present invention, excess sulfuric acid and/or
sulfur that is co-produced (e.g., beyond what can be used elsewhere
in the carbon capture and conversion process of certain embodiments
of the present invention) can be processed into a form suitable for
sale through process steps known in the art and science of
commercial sulfuric acid and/or sulfur production. Process heat can
also be generated in the production of hydrogen from hydrogen
sulfide. In certain embodiments of the present invention, process
heat generated in hydrogen production is recovered and utilized
elsewhere in the carbon capture and conversion process of certain
embodiments of the present invention to improve overall energy
efficiency. A chemical and/or heat and/or electrical co-product can
accompany the generation of molecular hydrogen for use as an
electron donor in certain embodiments of the present invention. The
chemical and/or heat and/or electrical co-products of molecular
hydrogen generation can be used to the extent possible elsewhere in
the carbon capture and conversion process of certain embodiments of
the present invention, for example, in order to improve efficiency
In certain embodiments, additional chemical co-product (e.g.,
beyond what can be used in the carbon capture and conversion
process of certain embodiments of the present invention) can be
prepared for sale in order to generate an additional stream of
revenue. Excess heat or electrical energy co-product in the
production of molecular hydrogen (e.g., beyond what can be used
internally in the process) can be delivered for sale, for example,
for use in another chemical and/or biological process through means
known in the art and science heat exchange and transfer and
electrical generation and transmission, including but not limited
to the conversion of process heat to electrical power in a form
that can be sold.
Certain embodiments of the present invention utilize
electrochemical energy stored in solid-state valence or conduction
electrons within an electrode or capacitor or related devices,
alone or in combination with chemical electron donors and/or
electron mediators to provide the oxyhydrogen microorganisms
reducing equivalents for the carbon-fixation reactions by means of
direct exposure of said electrode materials to the microbial
culturing environment and/or immersion of said electrode materials
within the microbial culture medium.
A feature of certain embodiments of the present invention regards
the production, or recycling of electron donors generated from
mineralogical origin that may also be used by certain oxyhydrogen
microbes as a source of reducing equivalents in addition, or in
lieu of hydrogen, including but not limited to electron donors
generated from reduced S and Fe containing minerals. Hence the
present invention, in certain embodiments, can enable the use of a
largely untapped source of energy--inorganic geochemical
energy.
The electron donors used in certain embodiments of the present
invention may be refined from natural mineralogical sources which
include but are not limited to one or more of the following:
elemental Fe.sup.0; siderite (FeCO.sub.3); magnetite
(Fe.sub.3O.sub.4); pyrite or marcasite (FeS.sub.2), pyrrhotite
(Fe.sub.(1-x)S (x=0 to 0.2)), pentlandite (Fe,Ni).sub.9S.sub.8,
violarite (Ni.sub.2FeS.sub.4), bravoite (Ni,Fe)S.sub.2,
arsenopyrite (FeAsS), or other iron sulfides; realgar (AsS);
orpiment (As.sub.2S.sub.3); cobaltite (CoAsS); rhodochrosite
(MnCO.sub.3); chalcopyrite (CuFeS.sub.2), bornite
(Cu.sub.5FeS.sub.4), covellite (CuS), tetrahedrite
(Cu.sub.8Sb.sub.2S.sub.7), enargite (Cu.sub.3AsS.sub.4), tennantite
(Cu.sub.12As.sub.4S.sub.13), chalcocite (Cu.sub.2S), or other
copper sulfides; sphalerite (ZnS), marmatite (ZnS), or other zinc
sulfides; galena (PbS), geocronite (Pb.sub.5(Sb,As.sub.2)S.sub.8),
or other lead sulfides; argentite or acanthite (Ag.sub.2S);
molybdenite (MoS.sub.2); millerite (NiS), polydymite
(Ni.sub.3S.sub.4) or other nickel sulfides; antimonite
(Sb.sub.2S.sub.3); Ga.sub.2S.sub.3; CuSe; cooperite (PtS); laurite
(RuS.sub.2); braggite (Pt,Pd,Ni)S; FeCl.sub.2.
The generation of electron donor from natural mineralogical sources
includes a preprocessing step in certain embodiments of the present
invention which can include but is not limited to comminuting,
crushing or grinding mineral ore to increase the surface area for
leaching with equipment such as a ball mill and wetting the mineral
ore to make a slurry. In these embodiments of the present invention
where electron donors are generated from natural mineral sources,
it may be advantageous if particle size is controlled so that the
sulfide and/or other reducing agents present in the ore may be
concentrated by methods known to the art including but not limited
to: flotation methods such as dissolved air flotation or froth
flotation using flotation columns or mechanical flotation cells;
gravity separation; magnetic separation; heavy media separation;
selective agglomeration; water separation; or fractional
distillation. After the production of crushed ore or slurry, the
particulate matter in the leachate or concentrate may be separated
by filtering (e.g. vacuum filtering), settling, or other well known
techniques of solid/liquid separation, prior to introducing the
electron donor containing solution to the chemoautotrophic culture
environment. In addition anything toxic to the chemoautotrophs that
is leached from the mineral ore may be removed prior to exposing
the chemoautotrophs to the leachate. The solid left after
processing the mineral ore may be concentrated with a filter press,
disposed of, retained for further processing, or sold depending
upon the mineral ore used in the particular embodiment of the
invention.
The electron donors in certain embodiments of the present invention
may also be refined from pollutants or waste products including but
not limited to one or more of the following: process gas; tail gas;
enhanced oil recovery vent gas; biogas; acid mine drainage;
landfill leachate; landfill gas; geothermal gas; geothermal sludge
or brine; metal contaminants; gangue; tailings; sulfides;
disulfides; mercaptans including but not limited to methyl and
dimethyl mercaptan, ethyl mercaptan; carbonyl sulfide; carbon
disulfide; alkanesulfonates; dialkyl sulfides; thiosulfate;
thiofurans; thiocyanates; isothiocyanates; thioureas; thiols;
thiophenols; thioethers; thiophene; dibenzothiophene;
tetrathionate; dithionite; thionate; dialkyl disulfides; sulfones;
sulfoxides; sulfolanes; sulfonic acid; dimethylsulfoniopropionate;
sulfonic esters; hydrogen sulfide; sulfate esters; organic sulfur;
sulfur dioxide and all other sour gases.
In addition to mineralogical sources, electron donors are produced
or recycled in certain embodiments of the present invention through
chemical reactions with hydrocarbons that may be of fossil origin,
but which are used in chemical reactions producing low or zero
carbon dioxide gas emissions. These reactions include
thermochemical and electrochemical processes. Such chemical
reactions that are used in these embodiments of the present
invention include but are not limited to: the thermochemical
reduction of sulfate reaction or TSR and the Muller-Kuhne reaction;
methane reforming-like reactions utilizing metal oxides in place of
water such as but not limited to iron oxide, calcium oxide, or
magnesium oxide whereby the hydrocarbon is reacted to form solid
carbonate with little or no emissions of carbon dioxide gas along
with hydrogen electron donor product.
Examples of reactions between metal oxides and hydrocarbons to
produce a hydrogen electron donor product and carbonates include
but are not limited to:
2CH.sub.4+Fe.sub.2O.sub.3+3H.sub.2O2FeCO.sub.3+7H.sub.2 and/or
CH.sub.4+CaO+2H.sub.2OCaCO.sub.3+4H.sub.2.
In certain embodiments, the generated electron donors are oxidized
in the chemosynthetic reaction step or steps by electron acceptors
that include but are not limited to carbon dioxide, oxygen and/or
one or more of the following: ferric iron or other transition metal
ions, nitrates, nitrites, sulfates, or valence or conduction band
holes in solid state electrode materials.
The position of the chemosynthetic and/or oxyhydrogen reaction step
or steps in the general process flow of certain embodiments of the
present invention is illustrated in FIG. 1 by Box 4 labeled
"Bioreactor--Knallgas Microbes."
At each step in the process where chemosynthetic and/or oxyhydrogen
reactions occur one or more types of electron donor and one or more
types of electron acceptor may be pumped or otherwise added to the
reaction vessel as either a bolus addition, or periodically, or
continuously to the nutrient medium containing oxyhydrogen
microorganisms. The chemosynthetic reaction driven by the transfer
of electrons from electron donor to electron acceptor can fix
inorganic carbon dioxide into organic compounds and biomass.
In certain embodiments of the present invention electron mediators
may be included in the nutrient medium to facilitate the delivery
of reducing equivalents from electron donors to oxyhydrogen
organisms in the presence of electron acceptors and inorganic
carbon in order to kinetically enhance the chemosynthetic reaction
step. This aspect of the present invention can be used to enhance
the transfer of reducing electrons to the oxyhydrogen microbes from
poorly soluble electron donors such as but not limited to H.sub.2
gas or electrons in solid state electrode materials using electron
mediators known in the art of electrical stimulation of microbial
metabolism including but not limited to
anthroquinone-2,6-disulfonate (AQDS), cobalt sepulchrate,
cytochromes, formate, humic substances, iron, methyl-viologen,
NAD+/NADH, neutral red (NR), phenazines, and quinones.
The delivery of reducing equivalents from electron donors to the
oxyhydrogen microorganisms for the chemosynthetic reaction or
reactions can be kinetically and/or thermodynamically enhanced in
certain embodiments through means including but not limited to: the
introduction of hydrogen storage materials into the microbial
culture environment that can double as a solid support media for
microbial growth--bringing absorbed or adsorbed hydrogen electron
donors into close proximity with the hydrogen-oxidizing
chemoautotrophs and/or the introduction of electrode materials
(e.g., graphite, graphite felt, activated carbon, carbon
nanofibers, conductive polymers, steel, iron, copper, titanium,
lead, tin, palladium, platinum, platinum-coated titanium, other
platinum coated metals, transition metals, transition metal alloys,
transition metal sulfides, oxides, chalcogenides, halides,
hydroxides, oxyhydroxides, phosphates, sulfates, and/or carbonates)
that can double as a solid growth support media and a source of
electron donors or acceptors directly into the chemoautotrophic
culture environment--bringing solid state electrons into close
proximity with the microbes. Some such embodiments of the present
invention can be useful for transferring reducing equivalents from
poorly soluble electron donors such as but not limited to H.sub.2
gas or electrons in solid state electrode materials to the
oxyhydrogen microorganisms.
The culture broth used in the chemosynthetic steps of certain
embodiments of the present invention may be an aqueous solution
containing suitable minerals, salts, vitamins, cofactors, buffers,
and other components needed for microbial growth, known to those
skilled in the art [Bailey and Ollis, Biochemical Engineering
Fundamentals, 2nd ed; pp 383-384 and 620-622; McGraw-Hill: New York
(1986)]. These nutrients can be chosen to maximize carbon-fixation
and promote the carbon flow through enzymatic pathways leading to
desired organic compounds. Alternative growth environments such as
those used in the arts of solid state or non-aqueous fermentation
may be used in certain embodiments. In certain embodiments that
utilize an aqueous culture, broth, salt water, sea water and/or
water from other natural bodies of water, or other non-potable
sources of water may be used when tolerated by the oxyhydrogen
microorganisms.
The biochemical pathways may be controlled and optimized in certain
embodiments of the present invention for the production of chemical
products (e.g., targeted organic compounds) and/or biomass by
maintaining specific growth conditions (e.g., levels of nitrogen,
oxygen, phosphorous, sulfur, trace micronutrients such as inorganic
ions, and if present any regulatory molecules that might not
generally be considered a nutrient or energy source). Depending
upon the embodiment of the invention the broth may be maintained in
aerobic, microaerobic, anoxic, anaerobic, or facultative
conditions. A facultative environment is considered to be one
having aerobic upper layers and anaerobic lower layers caused by
stratification of the water column.
The oxygen level is controlled in certain embodiments of the
invention. The oxygen level can be controlled, for example, to
enhance the production of targeted organic compounds by the
oxyhydrogen microorganisms through carbon-fixation. One objective
of controlling oxygen levels, in certain embodiments, is to control
(e.g., optimize) the intracellular Adenosine Triphosphate (ATP)
concentration through the cellular reduction of oxygen and
production of ATP by oxidative phosphorylation. In some such
embodiments, it can be desirable, while controlling ATP
concentration, to simultaneously keep the environment sufficiently
reducing so that the intracellular ratio of NADH (or NADPH) to NAD
(or NADP) remains relatively high. In some embodiments, ATP levels
are increased and/or optimized within the oxyhydrogen
microorganisms by means including but not limited to one or more of
the following: the cellular reduction of oxygen and/or another
electron acceptor of sufficient oxidation strength for ATP
production through oxidative phosphorylation; the direct
introduction of ATP into the culture medium; and/or the direct
introduction of chemical analogues of ATP into the culture
medium.
The reduction of oxygen by hydrogen in the oxyhydrogen reaction is
generally enzymatically linked to the production of ATP through
oxidative phosphorylation in oxyhydrogen microorganisms. The
oxyhydrogen reaction can act as a proxy for the light reaction in
photosynthesis in generating both NADPH and ATP. Generally, in
oxyhydrogen microorganisms, hydrogenase catalyzes the reduction of
NAD to NADH by hydrogen (or, alternatively, in some photosynthetic
organisms that are capable of carrying out the oxhydrogen reaction,
a hydrogenase catalyzes the reduction of ferrodoxin by H.sub.2,
which in turn reduces NADP to NADPH) [Chen, Gibbs, Plant Physiol.
(1992) 100, 1361-1365]. NADH and/or NADPH can then be used as
reducing agents for anabolic reactions, or to generate ATP by
reducing oxygen through oxidative phosphorylation [Bongers, J.
Bacteriology, (October 1970) 145-151]. Therefore, in place of the
following light dependent photosynthetic reaction:
2H.sub.2O+2NADP.sup.++2ADP+2Pi+light2NADPH+2H.sup.++2ATP+O.sub.2 an
oxyhydrogen reaction of
1/2O.sub.2+2NADP.sup.++2ADP+2Pi+3H.sub.22NADPH+2H.sup.++2ATP+2H.sub.2O
can occur in dark conditions (e.g., in the substantial absence of
visible electromagnetic radiation), with hydrogen acting in the
place of photons given the production of 2ATP per H.sub.2 consumed
[Bongers, J. Bacteriology, (October 1970) 145-151].
The maintenance of high intracellular concentrations of ATP as well
as NADH and/or NADPII is targeted in certain embodiments of the
present invention to promote carbon fixation and drive anabolic
pathways and/or solventogenic pathways that consume reducing
equivalents and either consume ATP, and/or that lower the net ATP
yield of chemosynthetic carbon-fixation. Such biochemical pathways
include but are not limited to the following: fatty acid synthesis;
mevalonate pathway and terpenoid synthesis; butanol pathway and
1-butanol synthesis; acetolactate/alpha-ketovalerate pathway and
2-butanol synthesis; and the ethanol pathway. A preferred oxygen
level can be determined, in some embodiments of in the present
invention: too low an oxygen level can reduce the intracellular ATP
in oxyhydrogen microorganisms below a desired level, while too high
an oxygen level can decrease the NADII (or NADPII) to NAD (or NADP)
ratio below a desired level.
The application of the oxyhydrogen reaction for the production of
ATP and NADH and/or NADPH used for carbon fixation and synthesis of
organic compounds in certain embodiments of the present invention
can provide advantages over alternative approaches using, for
example, anaerobic biochemical pathways for carbon-fixation for
such as Wood-Ljungdahl or methanogenic pathways. Carbon-fixation
through the Wood-Ljungdahl or methanogenic pathways generally
produces C1 or C2 organic compounds and it can be difficult to
produce longer than C4 compounds through these pathways.
The Wood-Ljungdahl pathway can produce acetic acid, ethanol,
butyric acid, and butanol in nature, but butyric acid and butanol
are generally minor products of H.sub.2 and CO.sub.2 gas
fermentation, and chain lengths longer than C4 do not typically
arise [Lynd, Zeikus, J. of Bacteriology (1983) 1415-1423; Eichler,
Schink, Archives of Microbiology (1984) 140, 147-152]. The
acetogenic pathways to acetic acid and butyric acid produce net
ATP, while the solventogenic pathways to ethanol and butanol do not
[Papoutsakis, Biotechnology & Bioengineering (1984) 26,
174-187; IIeise, Muller, Gottschalk, J. of Bacteriology (1989)
5473-5478; Lee, Park, Jang, Nielsen, Kim, Jung, Biotechnology &
Bioengineering (2008) 101, 2, 209-228]. Since ATP is needed for
cell maintenance a certain amount of relatively undesirable
non-biofuel co-product (ie organic acids) from acetogens fixing
carbon through the Wood-Ljungdahl pathway will generally be present
which constitutes a waste of reducing equivalents and carbon.
The production of hydrocarbons with chain length longer than C4 is
most commonly accomplished biologically through fatty acid
biosynthesis [Fischer, Klein-Marcuschamer, Stephanolpoulos,
Metabolic Engineering (2008) 10, 295-304]. Unlike the solventogenic
pathways coming out of the Wood-Ljungdahl pathway, fatty acid
synthesis involves net ATP consumption. For example the following
gives the net reaction for synthesis of Palmitic acid (C16), in
this example starting from Acetyl-CoA:
8Acetyl-CoA+7ATP+II2O+14NADPII+14H.sup.+.fwdarw.Palmitic
acid+8CoA+14NADP.sup.++7ADP+7Pi
One difficulty with using anaerobic pathways such as Methanogenesis
or Wood-Ljungdahl for ATP production to drive fatty acid synthesis
is the ATP produced per II.sub.2 consumed is relatively low: one
ATP per 4H.sub.2 for methane [Thauer, R. K., Kaster, A. K.,
Seedorf, H., Buckel, W. & Hedderich, R. Methanogenic archaea:
ecologically relevant differences in energy conservation. Nat Rev
Microbiol 6, 579-591, doi:nrmicro1931 [pii]] or acetic acid
production and one ATP per 10II.sub.2 for butyric acid production
[Papoutsakis, Biotechnology & Bioengineering (1984) 26,
174-187; Heise, Muller, Gottschalk, J. of Bacteriology (1989)
5473-5478; Lee, Park, Jang, Nielsen, Kim, Jung, Biotechnology &
Bioengineering (2008) 101, 2, 209-228]. By contrast, for the
oxyhydrogen reaction, hydrogenotrophic oxyhydrogen microorganisms
can produce up to two ATP per H.sub.2 consumed [Bongers, J.
Bacteriology, (October 1970) 145-151]. In other words oxyhydrogen
microorganisms can produce up to eight times more ATP per H.sub.2
consumed than methanogenic or acetogenic microorganisms.
Furthermore the path to ATP production through the oxyhydrogen
reaction produces water which can readily be incorporated into the
process stream rather than the relatively undesirable acetic acid
or butyric acid products of acidogenesis that can upset the system
pH and can rise to concentrations toxic to the organisms.
The highest energy density fuel that can be practically reached
naturally through the Wood-Ljungdahl pathway with inorganic carbon
input is generally ethanol at 30 MJ/kg, although butanol at 36.1
MJ/kg might be possible. Production of diesel fuels (46.2 MJ/kg) or
JP-8 aviation fuel (43.15 MJ/kg) can generally be difficult and is
generally less efficient utilizing anaerobic pathways such as
Wood-Ljungdahl due to the increased amount of H.sub.2 that needs to
be consumed in strictly anaerobic pathways per ATP produced, which
is needed for fatty acid synthesis. However these high density,
infrastructure compatible liquid fuels can be readily produced
through fatty synthesis pathways driven by ATP and NADII or NADPII
generated by the oxyhydrogen reaction.
Biomass lipid content and lipid biosynthetic pathway efficiency are
two factors that can affect the overall efficiency of certain
embodiments of the present invention for converting CO.sub.2 and
other C1 compounds to longer chain compounds (e.g.,
infrastructure-compatible fuels). The biomass lipid content can
determine the proportion of carbon and reducing equivalents
directed towards the synthesis of fuel products, as opposed to
other components of biomass. The lipid content can determine the
amount of energy input from the reducing equivalents that can be
captured in final fuel product. Likewise, the metabolic pathway
efficiency can determine the amount of reducing equivalents that
must be consumed in converting CO.sub.2 and hydrogen to lipid along
the lipid biosynthetic pathway. Many oxyhydrogen microorganisms
include species rich in lipid content and containing efficient
pathways from H.sub.2 and CO.sub.2 to lipid. Certain embodiments of
the present invention use species with high lipid contents such as
but not limited to Rhodococcus opacus which can have a lipid
content of over 70% [Gouda, M. K., Omar, S. H., Chekroud, Z. A.
& Nour Eldin, H. M. Bioremediation of kerosene I: A case study
in liquid media. Chemosphere 69, 1807-1814,
doi:S0045-6535(07)00738-2; Waltermann, M., Luftmann, H.,
Baumeister, D., Kalscheuer, R. & Steinbuchel, A. Rhodococcus
opacus strain PD630 as a new source of high-value single-cell oil?
Isolation and characterization of triacylglycerols and other
storage lipids. Microbiology 146 (Pt 5), 1143-1149 (2000).] and/or
species utilizing highly efficiency metabolic pathways such as but
not limited to the reverse tricarboxylic acid cycle [i.e. reverse
citric acid cycle] to fix carbon [Miura, A., Kameya, M., Arai, II.,
Ishii, M. & Igarashi, Y. A soluble NADH-dependent fumarate
reductase in the reductive tricarboxylic acid cycle of
Hydrogenobacter thermophilus TK-6. J Bacteriol 190, 7170-7177,
doi:JB.00747-08 [pii] 10.1128/JB.00747-08 (2008).; Shively, J. M.,
van Keulen, G. & Meijer, W. G. Something from almost nothing:
carbon dioxide fixation in chemoautotrophs. Annu Rev Microbiol 52,
191-230, doi: 10.1146/annurev.micro.52.1.191 (1998).]. In terms of
energy efficiency, the reverse tricarboxylic acid pathway can be a
relatively favorable pathway. The synthesis of palmitic acid from
H.sub.2 and CO.sub.2 is generally about 15% more efficient in terms
of reducing equivalents consumed than palmitic acid synthesis in
acetogens, due to the increased ATP output per reducing equivalent
consumed in the oxyhydrogen reaction by oxyhydrogen
microorganisms.
The source of inorganic carbon used in the chemosynthetic reaction
process steps of certain embodiments of the present invention
includes but is not limited to one or more of the following: a
carbon dioxide-containing gas stream that may be pure or a mixture;
liquefied CO.sub.2; dry ice; dissolved carbon dioxide, carbonate
ion, or bicarbonate ion in solutions including aqueous solutions
such as sea water; inorganic carbon in a solid form such as a
carbonate or bicarbonate minerals. Carbon dioxide and/or other
forms of inorganic carbon can be introduced to the nutrient medium
contained in reaction vessels either as a bolus addition,
periodically, or continuously at the steps in the process where
carbon-fixation occurs. Organic compounds containing only one
carbon atom that can be used in the synthetic reaction process
steps of certain embodiments of the present invention include but
are not limited to one or more of the following: carbon monoxide,
methane, methanol, formate, formic acid, and/or mixtures containing
C1 chemicals including but not limited to various syngas
compositions generated from various gasified or steam-reformed
fixed carbon feedstocks.
In certain embodiments, organic compounds containing only one
carbon atom and/or electron donors are generated through the
gasification and/or pyrolysis of biomass and/or other organic
matter (e.g., biomass and/or other organic matter from waste or low
value sources), and provided as a syngas to the culture of
oxyhydrogen microorganism, where the ratio of hydrogen to carbon
monoxide in the syngas may or may not be adjusted through means
such as the water gas shift reaction, prior to the syngas being
delivered to the microbial culture. In certain embodiments, organic
compounds containing only one carbon atom and/or electron donors
are generated through methane steam reforming from methane or
natural gas (e.g., stranded natural gas, or natural gas that would
be otherwise flared or released to the atmosphere), or biogas, or
landfill gas, and provided as a syngas to the culture of
oxyhydrogen microorganism, where the ratio of hydrogen to carbon
monoxide in the syngas may or may not be adjusted through means
such as the water gas shift reaction, prior to the syngas being
delivered to the microbial culture.
In certain embodiments of the present invention, carbon dioxide
containing flue gases are captured from the smoke stack at
temperature, pressure, and gas composition characteristic of the
untreated exhaust, and directed with minimal modification into the
reaction vessels where carbon-fixation occurs. In some embodiments
in which impurities harmful to chemoautotrophic organisms are not
present in the flue gas, modification of the flue gas upon entering
the reaction vessels can be limited to the compression needed to
pump the gas through the reactor system and/or the heat exchange
needed to lower the gas temperature to one suitable for the
microorganisms.
Oxyhydrogen microorganisms generally have an advantage over strict
anaerobic acetogenic or methanogenic microorganisms for carbon
capture applications due to the higher oxygen tolerance of
oxyhydrogen microorganisms. Since industrial flue gas is one
intended source of CO.sub.2 for certain embodiments of the present
invention, the relatively high oxygen tolerance of oxyhydrogen
microorganisms, as compared with obligately anaerobic methanogens
or acetogens, can allow the O.sub.2 content of 2-6% found in
typical fluegas to be tolerated.
In embodiments in which carbon dioxide bearing flue gas is
transported through a system for dissolving the carbon dioxide into
solution (such as is well known in the art of carbon capture), the
scrubbed flue gas, (which generally primarily includes inert gases
such as nitrogen), can be released into the atmosphere.
Gases in addition to carbon dioxide that are dissolved into
solution and fed to the culture broth or dissolved directly into
the culture broth in certain embodiments of the present invention
include gaseous electron donors (e.g., hydrogen gas), but in
certain embodiments of the present invention, may include other
electron donors such as but not limited to carbon monoxide and
other constituents of syngas, hydrogen sulfide, and/or other sour
gases. A controlled amount of oxygen can also be maintained in the
culture broth of some embodiments of the present invention, and in
certain embodiments, oxygen will be actively dissolved into
solution fed to the culture broth and/or directly dissolved into
the culture broth.
The dissolution of oxygen, carbon dioxide, and/or electron donor
gases such as but not limited to hydrogen and/or carbon monoxide
into solution can be achieved in some embodiments of the present
invention using a system of compressors, flowmeters, and/or flow
valves known to one skilled in the art of bioreactor scale
microbial culturing, which can be fed into one of more of the
following commonly used systems for pumping gas into solution:
sparging equipment; diffusers including but not limited to dome,
tubular, disc, or doughnut geometries; coarse or fine bubble
aerators; and/or venturi equipment. In certain embodiments of the
present invention, surface aeration may also be performed using
paddle aerators and the like. In certain embodiments of the present
invention, gas dissolution is enhanced by mechanical mixing with an
impeller and/or turbine. In some embodiments, hydraulic shear
devices can be used to reduce bubble size.
In certain embodiments of the present invention that require the
active pumping of air or oxygen into the culture broth in order to
maintain favorable (e.g., optimal) oxygenation levels, oxygen
bubbles are injected into the broth at a desirable (e.g., the
optimal) diameter for mixing and oxygen transfer. This has been
found to be 2 mm for certain embodiments [Environment Research
Journal May/June 1999 pgs. 307-315]. In certain aerobic embodiments
of the present invention, a process of shearing the oxygen bubbles
is used to achieve this bubble diameter as described in U.S. Pat.
No. 7,332,077. In some embodiments, bubbles have an average
diameter of no larger than 7.5 mm and slugging is avoided.
In certain embodiments of the present invention utilizing hydrogen
as electron donor, hydrogen gas is fed to the chemoautotrophic
culture vessel by bubbling it through the culture medium and/or by
diffusing it through a membrane that contacts the culture medium
and is impermeable to the culture medium. The latter method is
considered safer for many embodiments, and can be preferred since
hydrogen accumulating in the gas phase can create explosive
conditions (the range of explosive hydrogen concentrations in air
is 4 to 74.5% and can he avoided in certain embodiments of the
present invention). In some embodiments, the membrane is coated
with a biofilm of the oxyhydrogen microorganisms such that the
hydrogen must diffuse through the microorganism after passage
through the membrane.
Additional chemicals required or useful for the maintenance and
growth of oxyhydrogen microorganisms as known in the art can be
added to the culture broth of certain embodiments of the present
invention. These chemicals may include but are not limited to:
nitrogen sources such as ammonia, ammonium (e g ammonium chloride
(NH.sub.4Cl), ammonium sulfate ((NH.sub.4).sub.2SO.sub.4)), nitrate
(e.g. potassium nitrate (KNO.sub.3)), urea or an organic nitrogen
source; phosphate (e.g. disodium phosphate (Na.sub.2IIPO.sub.4),
potassium phosphate (KH.sub.2PO.sub.4), phosphoric acid
(H.sub.3PO.sub.4), potassium dithiophosphate
(K.sub.3PS.sub.2O.sub.2), potassium orthophosphate
(K.sub.3PO.sub.4), dipotassium phosphate (K.sub.2HPO.sub.4));
sulfate; yeast extract; chelated iron; potassium (e.g. potassium
phosphate (KH.sub.2PO.sub.4), potassium nitrate (KNO.sub.3),
potassium iodide (KI), potassium bromide (KBr)); and other
inorganic salts, minerals, and trace nutrients (e.g. sodium
chloride (NaCl), magnesium sulfate (MgSO.sub.4 7H.sub.2O) or
magnesium chloride (MgCl.sub.2), calcium chloride (CaCl.sub.2) or
calcium carbonate (CaCO.sub.3), manganese sulfate
(MnSO.sub.47II.sub.2O) or manganese chloride (MnCl.sub.2), ferric
chloride (FeCl.sub.3), ferrous sulfate (FeSO.sub.47H.sub.2O) or
ferrous chloride (FeCl.sub.24H.sub.2O), sodium bicarbonate
(NaHCO.sub.3) or sodium carbonate (Na.sub.2CO.sub.3), zinc sulfate
(ZnSO.sub.4) or zinc chloride (ZnCl.sub.2), ammonium molybdate
(NH.sub.4MoO.sub.4) or sodium molybdate
(Na.sub.2MoO.sub.42H.sub.2O), cuprous sulfate (CuSO.sub.4) or
copper chloride (CuCl.sub.22H.sub.2O), cobalt chloride
(CoCl.sub.26II.sub.2O), aluminum chloride (AlCl.sub.36II.sub.2O),
lithium chloride (LiCl), boric acid (II.sub.3BO.sub.3), nickel
chloride NiCl.sub.26II.sub.2O), tin chloride (SnCl.sub.2II.sub.2O),
barium chloride (BaCl.sub.22H.sub.2O), copper selenate
(CuSeO.sub.45H.sub.2O) or sodium selenite (Na.sub.2SeO.sub.3),
sodium metavanadate (NaVO.sub.3), chromium salts). In certain
embodiments the mineral salts medium (MSM) formulated by Schlegel
et al may be used [Thermophilic bacteria, Jakob Kristjansson,
Chapter 5, Section III, CRC Press, (1992)].
In certain embodiments, the concentrations of nutrient chemicals
(e.g., the electron donors and acceptors), are maintained at
favorable levels (e.g., as close as possible to their respective
optimal levels) for enhanced (e.g., maximum) carbon uptake and
fixation and/or production of organic compounds, which varies
depending upon the oxyhydrogen species utilized but is known or
determinable without undue experimentation to one of ordinary skill
in the art of culturing oxyhydrogen microorganisms.
Along with nutrient levels, the waste product levels, pII,
temperature, salinity, dissolved oxygen and carbon dioxide, gas and
liquid flow rates, agitation rate, and pressure in the microbial
culture environment are controlled in certain embodiments of the
present invention. The operating parameters affecting
carbon-fixation can be monitored with sensors (e.g. using a
dissolved oxygen probe and/or an oxidation-reduction probe to gauge
electron donor/acceptor concentrations) and can be controlled
either manually or automatically based upon feedback from sensors
through the use of equipment including but not limited to actuating
valves, pumps, and agitators. The temperature of the incoming broth
as well as incoming gases can be regulated by means such as but not
limited to heat exchangers.
The dissolution of gases and nutrients needed to maintain the
oxyhydrogen culture and promote carbon-fixation, as well as the
removal of inhibitory waste products, can be enhanced by agitation
of the culture broth. Oxyhydrogen microorganisms can carry out
carbon-fixation reactions throughout the volume of the reaction
vessel, which provides an advantage over other approaches including
those that employ photosynthetic organisms, which are surface area
limited due to the light requirements of photosynthesis. The use of
agitation can further enhance this advantage by distributing the
microorganisms, nutrients, optimal growth environment, and/or
CO.sub.2 as widely and evenly as possible throughout the reactor
volume so that production is enhanced (e.g., the reactor volume in
which carbon-fixation reactions occur at an optimal rate is
maximized).
Agitation of the culture broth in certain embodiments of the
present invention can be accomplished by equipment including but
not limited to: recirculation of broth from the bottom of the
container to the top via a recirculation conduit; sparging with
carbon dioxide, electron donor gas (e.g. H.sub.2), oxygen, and/or
air; and/or a mechanical mixer such as but not limited to an
impeller (100-1000 rpm) or turbine.
In certain embodiments of the present invention, the chemical
environment, oxyhydrogen microorganisms, electron donors, electron
acceptors, oxygen, pH, and/or temperature levels are varied either
spatially and/or temporally over a series of bioreactors in fluid
communication, such that a number of different carbon-fixation
reactions and/or biochemical pathways to organic compounds are
carried out sequentially or in parallel.
The nutrient medium containing oxyhydrogen microorganisms can be
removed from the bioreactors in certain embodiments of the present
invention partially or completely, periodically or continuously,
and can be replaced with fresh cell-free medium, for example, to
maintain the cell culture in an exponential growth phase, to
maintain the cell culture in a growth phase (exponential or
stationary) with enhanced (e.g., optimal) carbon-fixation rates, to
replenish the depleted nutrients in the growth medium, and/or
remove inhibitory waste products.
The high growth rate attainable by oxyhydrogen species can allow
them to match or surpass the highest rates of carbon fixation
and/or biomass production per standing unit biomass that can be
achieved by photosynthetic microbes. Consequently, in certain
embodiments, surplus biomass can be produced. Surplus growth of
cell mass can be removed from the system to produce a biomass
product. In some embodiments, surplus growth of cell mass can be
removed from the system in order to maintain a desirable (e.g., an
optimal) microbial population and cell density in the microbial
culture for continued high carbon capture and fixation rates.
Another advantage of certain embodiments of the present invention
relates to the vessels used to contain the carbon-fixation reaction
environment and culture in the carbon capture and fixation process.
Exemplary culture vessels that can be used in some embodiments of
the present invention to culture and grow the oxyhydrogen
microorganisms for carbon dioxide capture and fixation include
those that are known to those of ordinary skill in the art of large
scale microbial culturing. Such culture vessels, which may be of
natural or artificial origin, include but are not limited to:
airlift reactors; biological scrubber columns; bioreactors; bubble
columns; caverns; caves; cisterns; continuous stirred tank
reactors; counter-current, upflow, expanded-bed reactors; digesters
and in particular digester systems such as known in the prior arts
of sewage and waste water treatment or bioremediation; filters
including but not limited to trickling filters, rotating biological
contactor filters, rotating discs, soil filters; fluidized bed
reactors; gas lift fermenters; immobilized cell reactors; lagoons;
membrane biofilm reactors; microbial fuel cells; mine shafts;
pachuca tanks; packed-bed reactors; plug-flow reactors; ponds;
pools; quarries; reservoirs; static mixers; tanks; towers; trickle
bed reactors; vats; vertical shaft bioreactors; and wells. The
vessel base, siding, walls, lining, and/or top can be constructed
out of one or more materials including but not limited to bitumen,
cement, ceramics, clay, concrete, epoxy, fiberglass, glass,
macadam, plastics, sand, sealant, soil, steels or other metals and
their alloys, stone, tar, wood, and any combination thereof. In
certain embodiments of the present invention where the oxyhydrogen
microorganisms either require a corrosive growth environment and/or
produce corrosive chemicals through the carbon-fixation reaction,
corrosion resistant materials can be used to line the interior of
the container contacting the growth medium.
Since oxyhydrogen microorganisms do not require sunlight in order
to fix CO.sub.2, they can be used in carbon capture and fixation
processes that avoid many of the shortcomings that can be
associated with photosynthetically based carbon capture and
conversion technologies. For example, the maintenance of
chemosynthesis does not require shallow, wide ponds, nor
bioreactors with high surface area to volume ratios and special
features like solar collectors or transparent materials. A
technology such as certain embodiments of the present invention
using oxyhydrogen microbes does not have the diurnal, geographical,
meteorological, or seasonal constraints typically associated with
photosynthetically based systems.
Certain embodiments of the present invention minimize material
costs by using chemosynthetic vessel geometries having a low
surface area to volume ratio, such as but not limited to cubic,
cylindrical shapes with medium aspect ratio, ellipsoidal or
"egg-shaped", hemispherical, or spherical shapes, unless material
costs are superseded by other design considerations (e.g. land
footprint size). The ability to use compact reactor geometries can
arise from the absence of a light requirement for chemosynthetic
reactions, in contrast to photosynthetic technologies where the
surface area to volume ratio must be large to provide sufficient
light exposure.
The oxyhydrogen microorganisms' lack of dependence on light also
can allow plant designs with a much smaller footprint than those
traditionally associated with photosynthetic approaches. For
example, in scenarios where the plant footprint needs to be
minimized due to restricted land availability, a long vertical
shaft bioreactor system can be used for chemosynthetic carbon
capture. A bioreactor of the long vertical shaft type is described,
for example, in U.S. Pat. Nos. 4,279,754, 5,645,726, 5,650,070, and
7,332,077.
Unless superseded by other considerations, certain embodiments of
the present invention minimize vessel surfaces across which high
losses of water, nutrients, and/or heat occur, and/or the
introduction of invasive predators into the reactor. The ability to
minimize such surfaces can arise from the lack of light
requirements for chemosynthesis. Photosynthetic based technologies
generally are not able to minimize such surfaces since surfaces
across which high losses of water, nutrients, and/or heat occur, as
well as losses due to predation are generally the same surfaces
across which the light energy necessary for photosynthesis is
transmitted.
The culture vessels of the present invention can, in some
embodiments, use reactor designs known to those of ordinary skill
in the art of large scale microbial culture to maintain an aerobic,
microaerobic, anoxic, anaerobic, or facultative environment
depending upon the embodiment of the present invention. For
example, similar to the design of many sewage treatment facilities,
in certain embodiments of the present invention, tanks are arranged
in a sequence, with serial forward fluid communication, where
certain tanks are maintained in aerobic conditions and others are
maintained in anaerobic conditions, in order to perform multiple
chemosynthetic, and in certain embodiments, heterotrophic,
processing steps on the carbon dioxide waste stream.
In certain embodiments of the present invention, the oxyhydrogen
microorganisms are immobilized within their growth environment
Immobilization of the microorganisms can be accomplished using any
media known in the art of microbial culturing to support
colonization by microorganisms including but not limited to growing
the microorganisms on a matrix, mesh, or membrane made from any of
a wide range of natural and synthetic materials and polymers
including but not limited to one or more of the following: glass
wool, clay, concrete, wood fiber, inorganic oxides such as
ZrO.sub.2, Sb.sub.2O.sub.3, or Al.sub.2O.sub.3, the organic polymer
polysulfone, or open-pore polyurethane foam having high specific
surface area. The microorganisms in certain embodiments of the
present invention may also be grown on the surfaces of unattached
objects distributed throughout the growth container as are known in
the art of microbial culturing that include but are not limited to
one or more of the following: beads; sand; silicates; sepiolite;
glass; ceramics; small diameter plastic discs, spheres, tubes,
particles, or other shapes known in the art; shredded coconut
hulls; ground corn cobs; activated charcoal; granulated coal;
crushed coral; sponge balls; suspended media; bits of small
diameter rubber (elastomeric) polyethylene tubing; hanging strings
of porous fabric, Berl saddles, Raschig rings. The materials used
in the microbial support media may include hydrogen storage and/or
electrode materials in order to enhance the transfer of reducing
equivalents to the oxyhydrogen microorganisms. The electrode
materials that can be used include but are not limited to one or
more of the following: graphite, activated carbon, carbon
nanofibers, conductive polymers, steel, iron, copper, titanium,
lead, tin, palladium, platinum, transition metals, transition metal
alloys, transition metal sulfides, oxides, chalcogenides, halides,
hydroxides, oxyhydroxides, phosphates, sulfates, or carbonates. The
hydrogen storage materials that may be used in this application
include but are not limited to titanium, graphite, activated
carbon, carbon nanofibers, iron, copper, lead, tin, metal hydrides
including but not limited to TiFeII.sub.2, TiII.sub.2, VII.sub.2,
ZrII.sub.2, NiH, NbH.sub.2, PdH, and polymers known in the art of
hydrogen storage including but not limited to Metal Organic
Frameworks (MOF), and nanoporous polymeric materials. In certain
embodiments, the hydrogen storage material does not react strongly
with water or have a strong or rapid effect on the pII of the
culture medium.
Inoculation of the oxyhydrogen culture into the culture vessel can
be performed by methods including but not limited to transfer of
culture from an existing oxyhydrogen culture inhabiting another
carbon capture and fixation system of certain embodiments of the
present invention and/or incubation from a seed stock raised in an
incubator. The seed stock of oxyhydrogen strains can be transported
and stored in forms including but not limited to a powder, a
liquid, a frozen form, or a freeze-dried form as well as any other
suitable form, which may be readily recognized by one skilled in
the art. In certain embodiments in which a culture is established
in a very large reactor, growth and establishment of cultures can
be performed in progressively larger intermediate scale containers
prior to inoculation of the full scale vessel.
The position of the process step or steps for the separation of
cell mass from the process stream in the general process flow of
certain embodiments of the present invention is illustrated in FIG.
1 by Box 5, labeled "Cell Separation".
Separation of cell mass from liquid suspension can be performed by
methods known in the art of microbial culturing [Examples of cell
mass harvesting techniques are given in International Patent
Application No. WO08/00558, published Jan. 8, 1998; U.S. Pat. No.
5,807,722; U.S. Pat. No. 5,593,886 and U.S. Pat. No. 5,821,111.]
including but not limited to one or more of the following:
centrifugation; flocculation; flotation; filtration using a
membranous, hollow fiber, spiral wound, or ceramic filter system;
vacuum filtration; tangential flow filtration; clarification;
settling; hydrocyclone. In certain embodiments where the cell mass
is immobilized on a matrix, it can be harvested by methods
including but not limited to gravity sedimentation or filtration,
and separated from the growth substrate by liquid shear forces.
In certain embodiments of the present invention, if an excess of
cell mass has been removed from the culture, it can be recycled
back into the cell culture as indicated by the process arrow
labeled "Recycled Cell Mass" in FIG. 1., along with fresh broth
such that sufficient biomass is retained in the chemosynthetic
reaction step or steps. This can allow for continued enhanced
(e.g., optimal) autotrophic carbon-fixation and production of
organic compounds. The cell mass recovered by the harvesting system
can be recycled back into the culture vessel, for example, using an
airlift or geyser pump. In certain embodiments, the cell mass
recycled back into the culture vessel is not exposed to
flocculating agents, unless those agents are non-toxic to the
microorganisms.
In certain embodiments of the present invention, the microbial
culture and carbon-fixation reaction is maintained using continuous
influx and removal of nutrient medium and/or biomass, in steady
state where the cell population and environmental parameters (e.g.
cell density, chemical concentrations) are targeted at a constant
(e.g., optimal) level over time. Cell densities can be monitored in
certain embodiments of the present invention by direct sampling, by
a correlation of optical density to cell density, and/or with a
particle size analyzer. The hydraulic and biomass retention times
can be decoupled so as to allow independent control of both the
broth chemistry and the cell density. Dilution rates can be kept
high enough so that the hydraulic retention time is relatively low
compared to the biomass retention time, resulting in a highly
replenished broth for cell growth. Dilution rates can be set at an
optimal trade-off between culture broth replenishment, and
increased process costs from pumping, increased inputs, and other
demands that rise with dilution rates.
To assist in the processing of the biomass product into biofuels or
other useful products, the surplus microbial cells in certain
embodiments of the invention can be broken open following the cell
recycling step using, for example, methods including but not
limited to ball milling, cavitation pressure, sonication, or
mechanical shearing.
The harvested biomass in some embodiments can be dried in the
process step or steps of Box 7, labeled "Dryer" in the general
process flow of certain embodiments of the present invention
illustrated in FIG. 1.
Surplus biomass drying can be performed in certain embodiments of
the present invention using technologies including but not limited
to centrifugation, drum drying, evaporation, freeze drying,
heating, spray drying, vacuum drying, and/or vacuum filtration.
Heat waste from the industrial source of flue gas can be used in
drying the biomass, in certain embodiments. In addition, the
chemosynthetic oxidation of electron donors is generally exothermic
and generally produces waste heat. In certain embodiments of the
present invention waste heat can be used in drying the biomass.
In certain embodiments of the invention, the biomass is further
processed following drying to aid the production of biofuels or
other useful chemicals through the separation of the lipid content
or other targeted biochemicals from the microbial biomass. The
separation of the lipids can be performed by using nonpolar
solvents to extract the lipids such as, but not limited to, hexane,
cyclohexane, ethyl ether, alcohol (isopropanol, ethanol, etc.),
tributyl phosphate, supercritical carbon dioxide, trioctylphosphine
oxide, secondary and tertiary amines, or propane. Other useful
biochemicals can be extracted using solvents including but not
limited to: chloroform, acetone, ethyl acetate, and
tetrachloroethylene.
The extracted lipid content of the biomass can be processed using
methods known in the art and science of biomass refining including
but not limited to one or more of the following--catalytic cracking
and reforming; decarboxylation; hydrotreatment; isomerization--to
produce petroleum and petrochemical replacements, including but not
limited to one or more of the following: JP-8 jet fuel, diesel,
gasoline, and other alkanes, olefins and aromatics. In some
embodiments, the extracted lipid content of the biomass can be
converted to ester-based fuels, such as biodiesel (fatty acid
methyl ester or fatty acid ethyl ester), through processes known in
the art and science of biomass refining including but not limited
to transesterification and esterification.
The broth left over following the removal of cell mass can be
pumped to a system for removal of the chemical products of
chemosynthesis and/or spent nutrients which are recycled or
recovered to the extent possible and/or disposed of.
The position of the process step or steps for the recovery of
chemical products from the process stream in the general process
flow of certain embodiments of the present invention is illustrated
in FIG. 1 by Box 8, labeled "Separation of chemical
co-products."
Recovery and/or recycling of chemosynthetic chemical products
and/or spent nutrients from the aqueous broth solution can be
accomplished in certain embodiments of the present invention using
equipment and techniques known in the art of process engineering,
and targeted towards the chemical products of particular
embodiments of the present invention, including but not limited to:
solvent extraction; water extraction; distillation; fractional
distillation; cementation; chemical precipitation; alkaline
solution absorption; absorption or adsorption on activated carbon,
ion-exchange resin or molecular sieve; modification of the solution
pII and/or oxidation-reduction potential, evaporators, fractional
crystallizers, solid/liquid separators, nanofiltration, and all
combinations thereof.
In certain embodiments of the present invention, free fatty acids,
lipids, or other medium or long chain organic compounds appropriate
for refinement to biofuel products that have been produced through
chemosynthesis can be recovered from the process stream at the step
at Box 8 in FIG. 1. These free organic molecules can be released
into the process stream solution from the oxyhydrogen
microorganisms through means including but not limited to cellular
excretion or secretion or cell lysis. In certain embodiments of the
present invention, the recovered organic compounds are processed
using methods known in the art and science of biomass refining
including but not limited to one or more of the following:
catalytic cracking and reforming; decarboxylation; hydrotreatment;
isomerization. Such processes can be used to produce petroleum and
petrochemical replacements, including but not limited to one or
more of the following: JP-8 jet fuel, diesel, gasoline, and other
alkanes, olefins and aromatics. Recovered fatty acids can be
converted to ester-based fuels, such as biodiesel (fatty acid
methyl ester or fatty acid ethyl ester), through processes known in
the art and science of biomass refining including but not limited
to transesterification and esterification.
In some embodiments, following the recovery of chemical products
from the process stream, the removal of the waste products is
performed as indicated by Box 9, labeled "Waste removal" in FIG. 1.
The remaining broth can be returned to the culture vessel along
with replacement water and/or nutrients.
In certain embodiments of the present invention involving
chemoautotrophic oxidization of electron donors extracted from a
mineral ore, a solution of oxidized metal cations can remain
following the chemosynthetic reaction steps. A solution rich in
dissolved metal cations can also result from a particularly dirty
flue gas input to the process such as from a coal fired plant. In
some such embodiments of the present invention, the process stream
can be stripped of metal cations by methods including but not
limited to: cementation on scrap iron, steel wool, copper or zinc
dust; chemical precipitation as a sulfide or hydroxide precipitate;
electrowinning to plate a specific metal; absorption on activated
carbon or an ion-exchange resin, modification of the solution pH
and/or oxidation-reduction potential, solvent extraction. In
certain embodiments of the present invention, the recovered metals
can be sold for an additional stream of revenue.
In certain embodiments, the chemicals that are used in processes
for the recovery of chemical products, the recycling of nutrients
and water, and the removal of waste have low toxicity for humans,
and if exposed to the process stream that is recycled back into the
growth container, low toxicity for the oxyhydrogen microorganisms
being used.
In certain embodiments of the present invention, the pH of the
microbial culture is controlled. To address a decrease in pH, a
neutralization step can be performed prior to recycling the broth
back into the culture vessel in order to maintain the pH within an
optimal range for microbial maintenance and growth. Neutralization
of acid in the broth can be accomplished by the addition of bases
including but not limited to: limestone, lime, sodium hydroxide,
ammonia, caustic potash, magnesium oxide, iron oxide. In certain
embodiments, the base is produced from a carbon dioxide
emission-free source such as naturally occurring basic minerals
including but not limited to calcium oxide, magnesium oxide, iron
oxide, iron ore, olivine containing a metal oxide, serpentine
containing a metal oxide, ultramafic deposits containing metal
oxides, and underground basic saline aquifers. If limestone is used
for neutralization, then carbon dioxide will generally be released,
which can be directed back into the growth container for uptake by
chemosynthesis and/or sequestered in some other way, rather than
released into the atmosphere.
An additional feature of certain embodiments of the present
invention relates to the uses of organic compounds and/or biomass
produced through the chemosynthetic process step or steps of
certain embodiments of the present invention. Uses of the organic
compounds and/or biomass produced include but are not limited to:
the production of liquid fuels including but not limited to JP-8
jet fuel, diesel, gasoline, octane, biodiesel, butanol, ethanol,
propanol, isopropanol, propane, alkanes, olefins, aromatics, fatty
alcohols, fatty acid esters, alcohols; the production of organic
chemicals including but not limited to 1,3-propanediol,
1,3-butadiene, 1,4-butanediol, 3-hydroxypropionate,
7-ADCA/cephalosporin, .epsilon.-caprolactone,
.gamma.-valerolactone, acrylate, acrylic acid, adipic acid,
ascorbate, aspartate, ascorbic acid, aspartic acid, caprolactam,
carotenoids, citrate, citric acid, DHA, docetaxel, erythromycin,
ethylene, gamma butyrolactone, glutamate, glutamic acid, HPA,
hydroxybutyrate, isopentenol, isoprene, isoprenoids, itaconate,
itaconic acid, lactate, lactic acid, lanosterol, levulinic acid,
lycopene, lysine, malate, malonic acid, peptides, omega-3 DIIA,
omega fatty acids, paclitaxel, PHA, PHB, polyketides, polyols,
propylene, pyrrolidones, serine, sorbitol, statins, steroids,
succinate, terephthalate, terpenes, THF, rubber, wax esters,
polymers, commodity chemicals, industrial chemicals, specialty
chemicals, paraffin replacements, additives, nutritional
supplements, neutraceuticals, pharmaceuticals, pharmaceutical
intermediates, personal care products; as raw material and/or
feedstock for manufacturing or chemical processes; as feed stock
for alcohol or other biofuel fermentation and/or gasification and
liquefaction processes and/or other biofuel production processes
including but not limited to catalytic cracking, direct
liquefaction, Fisher Tropsch processes, hydrogenation, methanol
synthesis, pyrolysis, transesterification, or microbial syngas
conversions; as a biomass fuel for combustion in particular as a
fuel to be co-fired with fossil fuels; as sources of
pharmaceutical, medicinal or nutritional substances; as a carbon
source for large scale fermentations to produce various chemicals
including but not limited to commercial enzymes, antibiotics, amino
acids, vitamins, bioplastics, glycerol, or 1,3-propanediol; as a
nutrient source for the growth of other microbes or organisms; as
feed for animals including but not limited to cattle, sheep,
chickens, pigs, or fish; as feed stock for methane or biogas
production; as fertilizer; soil additives and soil stabilizers.
An additional feature of certain embodiments of the present
invention relates to the optimization of oxyhydrogen microorganisms
for carbon dioxide capture, carbon fixation into organic compounds,
and the production of other valuable chemical co-products. This
optimization can occur through methods known in the art of
artificial breeding including but not limited to accelerated
mutagenesis (e.g. using ultraviolet light or chemical treatments),
genetic engineering or modification, hybridization, synthetic
biology or traditional selective breeding. For certain embodiments
of the present invention utilizing a consortium of microorganisms,
the community can be enriched with desirable oxyhydrogen
microorganisms using methods known in the art of microbiology
through growth in the presence of targeted electron donors
including but not limited to hydrogen, acceptors including but not
limited to oxygen, and environmental conditions.
An additional feature of certain embodiments of the present
invention relates to modifying biochemical pathways in oxyhydrogen
microorganisms for the production of targeted organic compounds.
This modification can be accomplished by manipulating the growth
environment and/or through methods known in the art of artificial
breeding including but not limited to accelerated mutagenesis (e.g.
using ultraviolet light or chemical treatments), genetic
engineering or modification, hybridization, synthetic biology or
traditional selective breeding. The organic compounds produced
through the modification include but are not limited to one or more
of the following: biofuels including but not limited to JP-8 jet
fuel, diesel, gasoline, biodiesel, butanol, ethanol, long chain
hydrocarbons, lipids, fatty acids, pseudovegetable oil, and methane
produced from biological reactions in vivo; or organic compounds
and/or biomass optimized as a feedstock for biofuel and/or liquid
fuel production through chemical post-processing. These forms of
fuel can be used as renewable/alternate sources of energy with low
greenhouse gas emissions.
In order to give specific examples of the overall biological and
chemical process for using oxyhydrogen microorganisms to capture
CO.sub.2 and produce biomass and other useful co-products, a
process flow diagram describing a specific embodiment of the
present invention is now provided and described. This specific
example should not be construed as limiting the present invention
in any way and is provided for the sole purpose of
illustration.
FIG. 2 includes an exemplary process flow diagram illustrating one
embodiment of the present invention for the capture of CO.sub.2 by
oxyhydrogen microorganisms and the production of lipid rich
biomass, which is converted to JP-8 jet fuel. In this set of
embodiments, a carbon dioxide-rich flue gas is captured from an
emission source such as a power plant, refinery, or cement
producer. The flue gas can then be compressed and pumped into
cylindrical anaerobic digesters containing one or more oxyhydrogen
microorganisms such as but not limited to: purple non-sulfur
photosynthetic bacteria including but not limited to
Rhodopseudomonas palustris, Rhodopseudomonas capsulata,
Rhodopseudomonas viridis, Rhodopseudomonas sulfoviridis,
Rhodopseudomonas blastica, and other Rhodopseudomonas sp.
In some embodiments, Rhodopseudomonas capsulata can be used as the
oxyhydrogen microorganism, and, in some cases, a doubling time of 6
hours for chemoautotrophic growth on hydrogen can be achieved. See,
for example, Madigan, Gest, J. Bacteriology (1979) 524-530, which
is incorporated herein by reference. In some embodiments, the
microbial doubling time can be less than 6 hours, or shorter. In
some embodiments, the dry biomass concentration can be at least
about 3 g/l, at least about 4 g/l, or at least 5 g/l at steady
state. In some embodiments, the biomass lipid content in the
oxyhydrogen microorganism can be at least about 10%, at least about
20%, at least about 30%, at least about 35%, or at least about 40%.
For example, in some embodiments, Rhodopseudomonas palustris can be
used as the oxyhydrogen microorganism. See, for example, Carlozzi,
Pintucci, Piccardi, Buccioni, Minieri, Lambardi, Biotechnol. Lett.,
(2009) DOI 10.1007/s10529-009-0183-2, which is incorporated herein
by reference. In certain embodiments, the biomass lipid content of
the oxyhydrogen microorganisms is at least 40%; there is a steady
state bioreactor cell density of at least 5 g/liter in a continuous
process; the microbial doubling time is at most 6 hours; the
process achieves at least a 40% energy efficiency in converting
hydrogen into biomass; and/or at least 60% of the biomass energy
content is stored as lipid (which corresponds to about 40% biomass
lipid content by weight).
In the set of embodiments illustrated in FIG. 2, hydrogen electron
donor and oxygen and carbon dioxide electron acceptors are added
continuously to the growth broth along with other nutrients
required for chemosynthesis and culture maintenance and growth that
are pumped into the digester. In certain embodiments, the hydrogen
source is a carbon dioxide emission-free process. Exemplary carbon
dioxide emission-free processes include, for example, electrolytic
or thermochemical processes powered by energy technologies
including but not limited to photovoltaics, solar thermal, wind
power, hydroelectric, nuclear, geothermal, enhanced geothermal,
ocean thermal, ocean wave power, tidal power. In the set of
embodiments illustrated in FIG. 2, oxygen serves as an electron
acceptor in the chemosynthetic reaction for the intracellular
production of ATP through the oxyhydrogen reaction linked to
oxidative phosphorylation. The oxygen can originate from the flue
gas, it can be generated from the water-splitting reaction used to
produce the hydrogen, and/or it can be taken from air. In FIG. 2,
carbon dioxide from the flue gas serves as an electron acceptor for
the synthesis of organic compounds through biochemical pathways
utilizing the ATP produced through the oxyhydrogen reaction and
NADH and/or NADPH produced from the intracellular enzymatically
catalyzed reduction of NAD.sup.+ or NADP.sup.+ by H.sub.2. The
culture broth can be continuously removed from the digesters and
flowed through membrane filters to separate the cell mass from the
broth. The cell mass can then be recycled back into the digesters
and/or pumped to post-processing where lipid extraction is
performed according to methods known to those skilled in the art.
The lipids can then be converted to JP-8 jet fuel using methods
known to those skilled in the art of biomass refining (see, for
example, U.S. DOE Energy Efficiency & Renewable Energy Biomass
Program, "National Algal Biofuels Technology Roadmap", May 2010,
which is incorporated herein by reference in its entirety.
Cell-free broth which has passed through the cell mass removing
filters can then be subjected to any necessary additional waste
removal treatments which depends on the source of flue gas. The
remaining water and nutrients can then be pumped back into the
digesters.
Some of the Rhodopseudomonas species have extremely versatile
metabolisms, making them capable of photoautotrophic,
photoheterotrophic, heterotrophic, as well as chemoautotrophic
growth and the ability to live in both aerobic and anaerobic
environments [Madigan, Gest, J. Bacteriology (1979) 524-530]. In
certain embodiments of the present invention the heterotrophic
capability of Rhodopseudomonas sp. is exploited to further improve
the efficiency of energy and carbon conversion to lipid product.
The non-lipid biomass remainder following lipid extraction is
composed of primarily protein and carbohydrate. In certain
embodiments of the present invention, some of the carbohydrate
and/or protein remainder following lipid extraction is acid
hydrolyzed to simple sugars and/or amino acids, the acid is
neutralized, and the solution of simple sugars and/or amino acids
are fed to a second heterotrophic bioreactor containing
Rhodopseudomonas sp. that consumes the biomass input and produces
additional lipid product, as illustrated in FIG. 2.
The Rhodopseudomonas palustris genome has been sequenced by the DOE
Joint Genome Institute [Larimer et. al (2003) Nature Biotechnology
22, 55-61]. It is reported that its genetic system is particularly
amenable to modification. In one set of embodiments of the present
invention the carbon-fixation reaction or reactions are performed
by Rhodopseudomonas sp. that have been improved, optimized or
engineered for the improved fixation of carbon dioxide and/or other
forms of inorganic carbon and/or the improved production of organic
compounds through methods including but not limited to one or more
of the following: accelerated mutagenesis, genetic engineering or
modification, hybridization, synthetic biology or traditional
selective breeding.
FIG. 3 includes an exemplary schematic diagram of a bioreactor 300,
which can be used in certain embodiments. Bioreactor 300 can be
used, for example, as the reactor illustrated as Box 4 in FIG. 1
labeled "Bioreactor--Knallgas Microbes" and/or as the reactor
illustrated as Box 4 in FIG. 2 labeled "Bioreactor--Purple
non-sulfur bacterial." Bioreactor 300 illustrated in FIG. 3 can be
operated to take advantage of the low solubilities of hydrogen and
oxygen gas in water and avoids dangerous mixtures of hydrogen and
oxygen gas. In addition, the bioreactor can provide the oxyhydrogen
microorganisms with the oxygen and hydrogen needed for cellular
energy and carbon fixation, for example, by sparging, bubbling, or
diffusing oxygen or air up a vertical liquid column filled with
culture medium.
Bioreactor 300 includes a first column 302 and a second column 304.
In the set of embodiments illustrated in FIG. 3, oxygen is
introduced to first column 302 while hydrogen or syngas is
introduced to second column 304, although in other embodiments,
their order may be reversed. The oxygen and/or hydrogen and/or
syngas can be introduced to their respective columns by, for
example, sparging, bubbling, and/or diffusion such that they travel
upwards through the culture medium. Bioreactor 300 can include a
horizontal liquid connection 312 at the top of the columns and a
horizontal liquid connection 314 at the bottom of the columns.
In some embodiments, the level of the liquid medium with column 302
is maintained such that gaseous headspace 316 is formed above the
liquid. In addition, in some cases, the level of liquid medium
within column 304 can be arranged such that gaseous headspace 318
is formed above the liquid medium. In some embodiments, headspace
316 and/or headspace 318 can occupy at least about 2%, at least
about 10%, at least about 25%, between 2% and about 80%, between
about 10% and about 80%, or between about 25% and about 80% of the
total volume of the column in which they are positioned. Headspaces
316 and 318 can be isolated from each other by the liquid medium.
In some embodiments, the low solubility of the gases in the liquid
medium allow for the collection of gases at the tops of the columns
after bubbling or diffusing the gases up through their respective
columns. Establishing isolated headspaces can prevent a dangerous
amount of hydrogen and oxygen gases from mixing with each other.
For example, the hydrogen gas in one column can be prevented from
mixing with the oxygen gas in other column (and vice versa).
Inhibiting mixing of the hydrogen and oxygen gases can be achieved,
for example, by maintaining the connections between the two columns
such that they are filled with liquid, thereby preventing transport
of the gases from one column to the other. In some embodiments,
headspaces 316 and/or 318 can remain substantially stationary at
the top of their respective columns as liquid medium is circulated
between the first and second columns.
In FIG. 3, the horizontal liquid connection 312 at the top of the
columns and horizontal liquid connection 314 bottom of the columns
are arranged such that they allow the liquid medium to flow up one
column in the direction of the oxygen gas, and down the other
column, countercurrent to the hydrogen gas and/or syngas while the
horizontal liquid connections remain continuously filled. In other
embodiments, the liquid medium can flow up the column containing
the hydrogen gas and/or syngas and down the other column containing
the oxygen gas(in countercurrent flow relative to the gas).
In some embodiments, the gas on one side or the other, but not both
sides simultaneously, may be bubbled forcefully such that that
particular column acts as an airlift reactor and drives the
circulation of the culture medium between the two columns. In some
embodiments, the circulation of the fluid may also be assisted by
impellers, turbines, and/or pumps.
In some such embodiments, any unused hydrogen gas and/or syngas
that passes through the culture medium without being taken up by
the microorganisms (and which may end up in the head space) can be
recirculated by pumping the gas out of the head space, optionally
compressing it, and pumping it back into the medium at the bottom
of the liquid column on the hydrogen and/or syngas side. In some
embodiments, the oxygen and/or air might be similarly be
recirculated on its respective side or alternatively vented after
passing through the headspace.
The oxyhydrogen microorganisms are allowed to freely circulate
along with the liquid medium between the first and second columns
in certain embodiments. In other embodiments, the oxyhydrogen
microorganisms are restricted to the hydrogen side, for example, by
using a microfilter that retains the microorganisms on the hydrogen
side but allows the liquid medium to pass through.
FIG. 4 includes an exemplary schematic illustration of another
method of operating bioreactor 300 that can be used in certain
embodiments. The bioreactor arrangement in FIG. 4 can take
advantage of the relatively high solubility of carbon dioxide
and/or the strong ability of oxyhydrogen microorganism to capture
carbon dioxide from relatively dilute streams. The operation
illustrated in FIG. 4 can exploit the carbon concentrating
mechanism native to oxyhydrogen microorganisms. Flue gas and/or air
containing carbon dioxide can be transported through the oxygen
side of the bioreactor. The carbon dioxide can be dissolved into
solution and/or taken up by the oxyhydrogen microbes and
subsequently transported over to the hydrogen side of the reactor,
for example, through the horizontal liquid connection 312 at the
top of the column. On the hydrogen side, reducing equivalents can
be provided that drive fixation of the carbon. In some embodiments,
other gases pumped in on the oxygen side (e.g., oxygen, nitrogen,
etc.) have a low solubility relative to CO.sub.2, and are not
carried over to the hydrogen side. Rather than being passed from
column 302 to column 304, the low solubility gases can be
transported to headspace 316. In some embodiments, after the gases
are transported to headspace 316, they can be vented.
FIG. 5 includes an exemplary schematic diagram of an electrolysis
apparatus 500, which can be used in certain embodiments.
Electrolysis apparatus 500 can be used, for example, as the unit
illustrated as Box 3 in FIG. 1 labeled "Electron donor generation"
and/or as the unit illustrated as Box 3 in FIG. 2 labeled
"Electrolysis." Electrolysis apparatus 500 can be designed to take
advantage of the oxyhydrogen microorganisms' tolerance and need for
a certain concentration of oxygen by decreasing or eliminating the
complete separation of the hydrogen and oxygen produced from the
electrolysis step, relative to the separation schemes employed in
conventional electrolysis systems designed for the production of
pure hydrogen. Apparatus 500 includes an electrolysis unit 502 that
is configured to generate II.sub.2 and O.sub.2 from water. Any
suitable electrolysis unit 502 can be employed to perform the
electrolysis step. In some embodiments, the electrical resistance
in electrolysis unit 502 can be reduced at the expense of complete
hydrogen and oxygen separation by means including but not limited
one or more of the following: removing the separator used to
prevent gas crossover in standard electrolyzers and/or using a
relatively short distance between positive and negative
electrodes.
Apparatus 500 can include an outlet 504, through which the hydrogen
and oxygen produced by the electrolysis unit 502 can be
transported. Outlet 504 can be equipped with a separator 506, which
can be used to separate at least a portion of the hydrogen from at
least a portion of the oxygen. In certain embodiments,
semipermeable membranes such as polymer membranes designed for
H.sub.2 separation can be employed as separator 506. In certain
embodiments, separator 506 can include metal foils including but
not limited to foils made from palladium, palladium alloys,
vanadium, niobium, tantalum and their alloys, and/or other metals
and/or alloys that are permeable to hydrogen but less permeable to
other gases such as oxygen. In some embodiments, the separator can
be used to separate the hydrogen from the oxygen such that the
hydrogen content of one gas product exiting the separator is
enriched to a level that is desirable for oxyhydrogen microbes. The
gas product can then be transported to a bioreactor, where it can
be used as a feedstock. In certain embodiments, the amount of
hydrogen in one of the gas products exiting the separator can be
set at a level such that oxyhydrogen microorganism activity is
maximized, and the loss of hydrogen produced through electrolysis
apparatus 500 is minimized
The following documents are incorporated herein by reference in
their entirety for all purposes: U.S. Provisional Patent
Application No. 61/328,184, filed Apr. 27, 2010 and entitled "USE
OF OXYHYDROGEN MICROORGANISMS FOR NON-PHOTOSYNTHETIC CARBON CAPTURE
AND CONVERSION OF INORGANIC CARBON SOURCES INTO USEFUL ORGANIC
COMPOUNDS"; International Patent Application Serial No.
PCT/US2010/001402, filed May 12, 2010, entitled "BIOLOGICAL AND
CHEMICAL PROCESS UTILIZING CHEMOAUTOTROPHIC MICROORGNISMS FOR THE
CHEMOSYTHETIC FIXATION OF CARBON DIOXIDE AND/OR OTHER INORGANIC
CARBON SOURCES INTO ORGANIC COMPOUNDS, AND THE GENERATION OF
ADDITIONAL USEFUL PRODUCTS"; and U.S. Patent Application
Publication No. 2010/0120104, filed Nov. 6, 2009, entitled
"BIOLOGICAL AND CHEMICAL PROCESS UTILIZING CHEMOAUTOTROPHIC
MICROORGNISMS FOR THE CHEMOSYTHETIC FIXATION OF CARBON DIOXIDE
AND/OR OTHER INORGANIC CARBON SOURCES INTO ORGANIC COMPOUNDS, AND
THE GENERATION OF ADDITIONAL USEFUL PRODUCTS.
The following examples are intended to illustrate certain
embodiments of the present invention, but do not exemplify the full
scope of the invention.
EXAMPLE 1
In this example, oxyhydrogen microorganisms that accumulate high
lipid content and/or other valuable compounds such as
polyhydroxybutyrate (PIIB) to are grown on an inorganic medium with
CO.sub.2 as the carbon source and hydrogen acting as the electron
donor while oxygen provides the electron acceptor. Oxyhydrogen
microbes such as these can be used in certain embodiments of the
present invention in converting C1 chemicals such as carbon dioxide
into longer chain organic chemicals.
Static anaerobic reaction vessels were inoculated with Cupriavidus
necator DSM 531 (which can accumulate a high percentage of cell
mass as PHB). The inoculum were taken from DSM medium no. 1 agar
plates kept under aerobic conditions at 28 degrees Celsius. Each
anaerobic reaction vessel had 10 ml of liquid medium DSM no. 81
with 80% H.sub.2, 10% CO.sub.2 and 10% O.sub.2 in the headspace.
The cultures were incubated at 28 degrees Celsius. The Cupriavidus
necator reached an optical density (OD) at 600 nm of 0.98 and a
cell density of 4.7.times.10.sup.8 cells/ml after 8 days.
Another growth experiment was performed for Cupriavidus necator
(DSM 531). The medium used for growth was the mineral salts medium
(MSM) formulated by Schlegel et al. The MSM medium was formed by
mixing 1000 ml of Medium A, 10 ml of Medium B, and 10 ml of Medium
C. Medium A included 9 g/l Na.sub.2HPO.sub.412H.sub.2O, 1.5 g/l
KH.sub.2PO.sub.4, 1.0 g/l, 0.2 g/l MgSO.sub.47H.sub.2O, and 1.0 ml
of Trace Mineral Medium. The Trace Mineral Medium included 1000 ml
distilled water; 100 mg/l ZnSO.sub.4.7H.sub.2O; 30 mg/l
MnCl.sub.24H.sub.2O; 300 mg/l H.sub.3BO.sub.3; 200 mg/l
COCl.sub.26H.sub.2O; 10 mg/l CuCl.sub.22H.sub.2O; 20 mg/l
NiCl.sub.26H.sub.2O; and 30 mg/l Na.sub.2MoO.sub.42H.sub.2O. Medium
B contained 100 ml of distilled water; 50 mg ferric ammonium
citrate; and 100 mg CaCl.sub.2. Medium C contained 100 ml of
distilled water and 5 g NaHCO.sub.3.
The cultures were grown in 20 ml of MSM media in 150-ml stopped and
sealed serum vials with the following gas mixture in the headspace:
71% Hydrogen; 4% Oxygen; 16% Nitrogen; 9% Carbon dioxide. The
headspace pressure was 7 psi. The cultures were grown for eight
days at 30 degrees Celsius. Cupriavidus necator reached an OD at
600 nm of 0.86.
It is known that, in larger scale bioreactor equipment, faster
growth rates and higher cell densities can be attained.
Accordingly, it is believed that higher growth rates and cell
densities can be achieved simply by scaling up the systems
described above. For example Cupriavidus necator which is also
known as Alcaligenes eutrophus, Ralstonia eutropha, Hydrogenomona
eutropha, has been grown in bioreactors on H.sub.2/CO.sub.2/O.sub.2
to a cell density of over 90 grams/liter [Tanaka, Ishizaki;
Biotech. And Bioeng., vol. 45, 268-275 (1995)], and with doubling
times below two hours [Ammann, Reed, Durichek, Appl. Microbio.,
(1968) 822-826].
Specific preferred embodiments of the present invention have been
described here in sufficient detail to enable those skilled in the
art to practice the full scope of invention. However it is to be
understood that many possible variations of the present invention,
which have not been specifically described, still fall within the
scope of the present invention and the appended claims. Hence these
descriptions given herein are added only by way of example and are
not intended to limit, in any way, the scope of this invention.
More generally, those skilled in the art will readily appreciate
that all parameters, dimensions, materials, and configurations
described herein are meant to be exemplary and that the actual
parameters, dimensions, materials, and/or configurations will
depend upon the specific application or applications for which the
teachings of the present invention is/are used. Those skilled in
the art will recognize, or be able to ascertain using no more than
routine experimentation, many equivalents to the specific
embodiments of the invention described herein. It is, therefore, to
be understood that the foregoing embodiments are presented by way
of example only and that, within the scope of the appended claims
and equivalents thereto, the invention may be practiced otherwise
than as specifically described and claimed. The present invention
is directed to each individual feature, system, article, material,
kit, and/or method described herein. In addition, any combination
of two or more such features, systems, articles, materials, kits,
and/or methods, if such features, systems, articles, materials,
kits, and/or methods are not mutually inconsistent, is included
within the scope of the present invention.
The indefinite articles "a" and "an," as used herein in the
specification and in the claims, unless clearly indicated to the
contrary, should be understood to mean "at least one."
The phrase "and/or," as used herein in the specification and in the
claims, should be understood to mean "either or both" of the
elements so conjoined, i.e., elements that are conjunctively
present in some cases and disjunctively present in other cases.
Other elements may optionally be present other than the elements
specifically identified by the "and/or" clause, whether related or
unrelated to those elements specifically identified unless clearly
indicated to the contrary. Thus, as a non-limiting example, a
reference to "A and/or B," when used in conjunction with open-ended
language such as "comprising" can refer, in one embodiment, to A
without B (optionally including elements other than B); in another
embodiment, to B without A (optionally including elements other
than A); in yet another embodiment, to both A and B (optionally
including other elements); etc.
As used herein in the specification and in the claims, "or" should
be understood to have the same meaning as "and/or" as defined
above. For example, when separating items in a list, "or" or
"and/or" shall be interpreted as being inclusive, i.e., the
inclusion of at least one, but also including more than one, of a
number or list of elements, and, optionally, additional unlisted
items. Only terms clearly indicated to the contrary, such as "only
one of" or "exactly one of," or, when used in the claims,
"consisting of," will refer to the inclusion of exactly one element
of a number or list of elements. In general, the term "or" as used
herein shall only be interpreted as indicating exclusive
alternatives (i.e. "one or the other but not both") when preceded
by terms of exclusivity, such as "either," "one of," "only one of,"
or "exactly one of." "Consisting essentially of," when used in the
claims, shall have its ordinary meaning as used in the field of
patent law.
In the claims, as well as in the specification above, all
transitional phrases such as "comprising," "including," "carrying,"
"having," "containing," "involving," "holding," and the like are to
be understood to be open-ended, i.e., to mean including but not
limited to. Only the transitional phrases "consisting of" and
"consisting essentially of" shall be closed or semi-closed
transitional phrases, respectively.
* * * * *